Aminonorbornane derivative and manufacture method therefor and use thereof

ABSTRACT

The present invention relates to a compound of formula I or a pharmaceutically acceptable salt, a solvate, an active metabolite, a polymorph, an ester, an optical isomer or a prodrug thereof, a pharmaceutical composition comprising the compound of formula I, and use of the compound, as a Bruton&#39;s tyrosine kinase inhibitor with high selectivity for BTK(C481S) mutant, in the manufacture of a medicament for preventing or treating a heteroimmune disease, an autoimmune disease or a cancer.

TECHNICAL FIELD

The present invention belongs to the field of medicine, and particularlyrelates to an aminonorbornane derivative as a Bruton's tyrosine kinaseinhibitor with high selectivity for a C481S mutant, a pharmaceuticalcomposition thereof, a manufacture method therefor and use thereof inthe manufacture of a medicament.

BACKGROUND

B-cell receptor (BCR) signaling pathway plays a key role in thematuration, differentiation and development of B cells. AberrantBCR-mediated signal transduction may lead to deregulated B cellactivation and/or formation of pathogenic autoantibodies, resulting in avariety of human diseases including cancer, autoimmune diseasesincluding lupus erythematosus, chronic lymphocytic lymphoma, diffuselarge cell lymphoma, follicular lymphoma or chronic lymphocyticleukemia, heteroimmune diseases including inflammatory diseases, asthma,and the like.

Bruton's tyrosine kinase (BTK) is a member of the TEC family ofnon-receptor tyrosine kinases. It plays a key role in the activation ofthe BCR signaling pathway, and is a key regulator of early B-cellformation and mature B-cell activation and survival (Khan et al.,Immunity 1995 3:283; Ellmeier et al., J Exp Med 2000 192:1611). BTKplays an important role in regulating B-cell proliferation and apoptosis(Islam and Smith, Immmol Rev 2000 178:49; Davis et al., Nature 2010463:88-94), and therefore inhibition of BTK can be used to treat certainB-cell lymphomas and leukemia (Feldhahn et al., J Exp Med 2005201:1837).

The role of BTK in autoimmune and inflammatory diseases has beenconfirmed by BTK-deficient mouse models. In a preclinical mouse model ofsystemic lupus erythematosus (SLE), BTK-deficient mice show significantimprovement in progressive disease. In addition, BTK-deficient mice areresistant to collagen-induced arthritis (Jansson and Holmdahl, Clin ExpImmunol 1993 94:459). Selective BTK inhibitors show a clear dose-effectrelationship in mouse models of arthritis (Pan et al., Chem. Med. Chem.2007 2:58-61). Clinical studies are currently underway for the treatmentof arthritis with BTK inhibitors.

As the first marketed BTK inhibitor, ibrutinib (trade name Imbruvica)has had great success with its annual sales reaching 2.6 billion USdollars in 2017. However, as with many other anticancer drugs, somepatients exhibit resistance to the drug. C481S mutation of BTK kinasehas been found to be the main cause of drug resistance. Ibrutinib actspharmacodynamically by irreversible covalent binding to C481 tryptophanresidue of BTK kinase; however, the tryptophan residue loses the abilityto covalently binding to ibrutinib as C481S mutation changes tryptophaninto serine.

According to clinical statistics, BTK(C481S) mutation is associated with87% of patients with relapsed chronic lymphoma (CLL) (Woyach et al., JClin Oncol 2017 35:1437-1443) and about 80% of patients with relapsedmantle cell lymphoma (MCL) (Chiron et al., Cancer Discovery 2014 4(9):1-14). The development of a BTK inhibitor that is effective againstBTK(C481S) mutant would overcome the resistance to ibrutinib due to theC481S mutation.

SUMMARY

The technical problem to be solved by the prevent invention is toprovide a novel and undisclosed compound of Bruton's tyrosine kinaseinhibitor with high selectivity for BTK(C481S) mutant, apharmaceutically acceptable salt, a solvate, an active metabolite, apolymorph, an ester, an optical isomer or a prodrug thereof, use of thecompound in pharmacy and a method for preventing or treating diseasesrelated to excessive BTK activity in human or mammals by using thecompound disclosed herein.

In order to solve the technical problem, the technical scheme adopted inthe present invention is as follows:

Provided is a compound of formula (I)

or a pharmaceutically acceptable salt, a solvate, an active metabolite,a polymorph, an ester, an optical isomer or a prodrug thereof, wherein

ring A is selected from one of the following structures:

R⁵ being selected from hydrogen, halogen, cyano, hydroxyl, alkynyl,amino, C₁₋₃ alkyl, C₁₋₃ alkoxy, C₁₋₃ haloalkyl, C₁₋₃ haloalkoxy, C₁₋₃haloalkylamino, C₃₋₇ cycloalkyl, C₃₋₇ cycloalkoxy and C₃₋₇cycloalkylamino;

ring B is a substituted or unsubstituted aromatic ring or heteroaromaticring; ring C is a substituted or unsubstituted aromatic ring orheteroaromatic ring;

L is a single bond or one of the following structures:

R¹ is selected from R³ and one of the following structures:

R³ being selected from hydrogen, substituted or unsubstituted C₁₋₆alkyl, substituted or unsubstituted C₁₋₆ alkynyl, substituted orunsubstituted C₁₋₆ alkenyl, substituted or unsubstituted C₆₋₁₀ aryl,substituted or unsubstituted C₁₋₉ heteroaryl, substituted orunsubstituted C₃₋₇ cycloalkyl, and substituted or unsubstituted C₂₋₇heterocycloalkylamino; and

R⁴ being selected from hydrogen, substituted or unsubstituted C₁₋₆alkyl, substituted or unsubstituted C₆₋₁₀ aryl, substituted orunsubstituted C₁₋₉ heteroaryl, substituted or unsubstituted C₃₋₇cycloalkyl, and substituted or unsubstituted C₃₋₇ heterocycloalkyl; and

R² is selected from H, substituted or unsubstituted C₁₋₃ alkyl,substituted or unsubstituted C₃₋₇ cycloalkyl, substituted orunsubstituted C₂₋₇ heterocycloalkyl, substituted or unsubstituted C₆₋₁₀aryl, and substituted or unsubstituted C₁₋₉ heteroaryl.

R¹ and R², along with N attached thereto, form a substituted orunsubstituted C₂₋₇ heterocyclic ring, and R³ and R⁴, along with Nattached thereto, form or do not form a C₃₋₇ heterocyclylamino or a C₃₋₉heteroarylamino.

Preferably, for R³, a substituent of the substituted C₁₋₆ alkyl, thesubstituted C₁₋₆ alkynyl, the substituted C₁₋₆ alkenyl, the substitutedC₆₋₁₀ aryl, the substituted C₁₋₉ heteroaryl, the substituted C₃₋₇cycloalkyl or the substituted C₂₋₇ heterocycloalkyl is selected from oneor more of halogen, cyano, hydroxyl, amino, substituted or unsubstitutedacylamino, substituted or unsubstituted aminoacyl, substituted orunsubstituted C₁₋₄ alkyl, substituted or unsubstituted C₃₋₇ cycloalkyl,substituted or unsubstituted C₃₋₇ cycloalkoxy, substituted orunsubstituted C₁₋₄ alkylamino, di[substituted or unsubstituted C₁₋₄alkyl]amino, substituted or unsubstituted C₃₋₇ cycloalkylamino,substituted or unsubstituted C₃₋₇ heterocycloalkylamino, substituted orunsubstituted C₁₋₃ alkoxy, substituted or unsubstituted C₃₋₇cycloalkoxy, substituted or unsubstituted C₆₋₁₀ aryl, and substituted orunsubstituted C₃₋₇ heterocycloalkyl.

Preferably, for R⁴, a substituent of the substituted C₁₋₆ alkyl, thesubstituted C₆₋₁₀ aryl, the substituted C₁₋₉ heteroaryl, the substitutedC₃₋₇ cycloalkyl or the substituted C₃₋₇ heterocycloalkyl is selectedfrom one or more of halogen, hydroxyl, cyano, amino, substituted orunsubstituted C₁₋₄ alkenyl, substituted or unsubstituted C₃₋₇cycloalkyl, substituted or unsubstituted C₃₋₇ cycloalkoxy, substitutedor unsubstituted C₁₋₄ alkylamino, di[substituted or unsubstituted C₁₋₄alkyl]amino, substituted or unsubstituted C₃₋₇ cycloalkylamino,substituted or unsubstituted C₃₋₇ heterocycloalkylamino, substituted orunsubstituted C₁₋₃ alkoxy, substituted or unsubstituted C₃₋₇cycloalkoxy, substituted or unsubstituted C₆₋₁₀ aryl, and substituted orunsubstituted C₃₋₇ heterocycloalkyl.

Preferably, ring A is one of the following structures:

R⁵ being selected from hydrogen, halogen, cyano, hydroxyl, alkynyl,amino, C₁₋₃ alkyl, C₁₋₃ alkoxy, C₁₋₃ haloalkyl, C₁₋₃ haloalkoxy, C₁₋₃haloalkylamino. C₃₋₇ cycloalkyl, C₃₋₇ cycloalkoxy and C₃₋₇cycloalkylamino;

preferably, for ring B, a substituent of the substituted aromatic ringor heteroaromatic ring is selected from one or more of halogen,hydroxyl, cyano, amino, C₁₋₃ alkyl, C₁₋₃ alkoxy, C₁₋₃ haloalkyl and C₁₋₃haloalkoxy.

Preferably, ring B is one of the following structures:

Preferably, for ring C, a substituent of the substituted aromatic ringor heteroaromatic ring is selected from one or more of halogen,hydroxyl, cyano, amino. C₁₋₃ alkyl, C₁₋₃ alkoxy, C₁₋₃ haloalkyl and C₁₋₃haloalkoxy.

Preferably, ring C is one of the following structures:

Preferably, R² is selected from H, and R¹ is selected from

wherein R³ is selected from hydrogen, substituted or unsubstituted C₁₋₆alkyl, substituted or unsubstituted C₁₋₆ alkynyl, substituted orunsubstituted C₁₋₆ alkenyl, substituted or unsubstituted C₆₋₁₀ aryl,substituted or unsubstituted C₁₋₉ heteroaryl, substituted orunsubstituted C₃₋₇ cycloalkyl, and substituted or unsubstituted C₂₋₇heterocycloalkylamino.

Most preferably, the compound is selected from any one of the followingstructures:

Provided is use of the compound according to any one of aspects above inthe manufacture of a medicament for preventing or treating aheteroimmune disease, an autoimmune disease or a cancer,

wherein the heteroimmune disease, the autoimmune disease or the canceris associated with excessive activity of Bruton's tyrosine kinase, or

the heteroimmune disease, the autoimmune disease or the cancer isassociated with aberrant B-cell proliferation.

Further, the heteroimmune disease is an inflammatory disease or asthma.

Further, the autoimmune disease is lupus erythematosus, chroniclymphocytic lymphoma, diffuse large cell lymphoma, follicular lymphomaor chronic lymphocytic leukemia.

Provided is a pharmaceutical composition comprising one or morecompounds according to any one of aspects above.

Provided is a pharmaceutical formulation comprising a therapeuticallyeffective amount of the compound according to any one of aspects aboveand a pharmaceutically acceptable excipient.

The pharmaceutical formulation is formulated for oral administration,parenteral administration, buccal administration, nasal administration,topical administration or rectal administration.

The pharmaceutical formulation is for use in treating a disease orcondition associated with excessive activity of Bruton's tyrosinekinase, comprising administering the pharmaceutical formulation to ahuman or mammal in need thereof; the disease associated with excessiveactivity of Bruton's tyrosine kinase is a heteroimmune disease, anautoimmune disease or a cancer; the heteroimmune disease is aninflammatory disease or asthma; the autoimmune disease is lupuserythematosus, chronic lymphocytic lymphoma, diffuse large celllymphoma, follicular lymphoma or chronic lymphocytic leukemia.

The present invention comprises the step of contacting thepharmaceutical formulation with BTK, comprising an in vitro or in vivoassay.

Manufacture method I for the compound I above comprises: (S1) performingSuzuki coupling of compound IIIA with boronic acid or borate II to givecompound IV; (S2) converting the compound IV into the hydrochloride ofcompound V by removal of benzyloxycarbonyl with trifluoroacetic acid;and (S3) coupling the compound V with an organic acid to give thecompound I described above;

wherein X=halogen, and R², R³, L, ring A, ring B and ring C aredescribed as above.

Manufacture method 2 for the compound I above comprises: (A1) convertingcompound IIIA into the hydrochloride of compound VI by removal ofbenzyloxycarbonyl with trifluoroacetic acid; (A2) coupling the compoundVI with an organic acid to give compound VII; and (A3) performing Suzukicoupling of the compound VII with boronic acid or borate II to give thecompound I described above;

wherein X=halogen, and R², R, L, ring A, ring B and ring C are describedas above.

Manufacture method 3 for the compound I above comprises: (B1) performingChan-Lam-Evans coupling of compound IIIB with boronic acid II in thepresence of catalyzation of copper acetate to give compound VIII; (B2)converting the compound VIII into the hydrochloride of compound IX byremoval of benzyloxycarbonyl with trifluoroacetic acid; and (B3)coupling the compound IX with an organic acid to give the compound Idescribed above:

wherein R², R, L, ring A, ring B and ring C are described as above.

Each of the products resulting from the reactions in methods 1, 2 and 3may be obtained by conventional separation techniques, including but notlimited to filtration, distillation, crystallization, chromatographicseparation, and the like. The starting materials required for synthesismay be synthesized by oneself or purchased from commercialestablishments, such as Adrich or Sigma. These materials can becharacterized using conventional means, such as physical constants andspectral data. The compounds described herein may be synthesized to givea single optical isomer or a mixture of optical isomers.

The superscripts of letters in the present invention indicate thesequence number of the group, and the subscripts indicate the number ofthe atom. For example: R¹, R² and R³ represent the 1^(st) to the 3^(rd)R groups, and C₁₋₄ alkyl represents alkyl containing 1-4 C atoms. Thenumber of C atoms on the substituent is not counted in the main chain.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is an OCI-LY10 xenograft tumor model.

FIG. 2 is a TMD-8 xenograft tumor model.

DETAILED DESCRIPTION

The present invention can be better understood according to thefollowing examples.

How-ever, it is easily understood by those skilled in the art that thecontent described in the examples are only used to illustrate thepresent invention, and should not and will not limit the presentinvention described in detail in the claims.

Synthesis of Intermediate I-5

Synthetic Route of Intermediate I-5

I-2: methyl4-(((benzyloxy)carbonyl)amino)bicyclo[2.2.1]heptane-1-carboxylate

To a solution of 4-(methoxycarbonyl)bicyclo[2.2.1]heptane-1-carboxylicacid (1-1) (3.5 g, 17.7 mmol) and TEA (1.78 g, 17.7 mmol) in toluene (30mL) was added DPPA (5.34 g, 19.5 mmol). The mixture was heated to 90° C.and maintained at this temperature for 2 h. After being cooled to roomtemperature, the mixture was added with BnOH (1.9 g, 17.7 mmol). Theresulting mixture was stirred at 90° C. for 4 days. After being cooledto room temperature, the mixture was diluted with ethyl acetate andwashed with aqueous NaHCO₃ solution. The organic phase was separated,dried over Na₂SO₄, filtered and evaporated. The residue was purified bycolumn chromatography (ethyl acetate/petroleum ether=1:4) to give thedesired compound I-2 (3 g, yield: 56%).

I-3: 4-(((benzyloxy)carbonyl)amino)bicyclo[2.2.1]heptane-1-carboxylicacid

To a solution of methyl4-(((benzyloxy)carbonyl)amino)bicyclo[2.2.1]heptane-1-carboxylate(I-2)(3 g, 9.9 mmol) was added NaOH (792 mg, 19.8 mmol), and the mixturewas heated to 60° C. and maintained at this temperature for 10 h. Themixture was concentrated, added with water (50 mL), and then added with1N aqueous HCl solution to adjust the pH to 4. Ethyl acetate (20 mL×3)was added for extraction, and the organic phase was separated, driedover Na₂SO₄, filtered and evaporated to give the desired compound I-3(2.2 g, yield: 77%), which was directly used in the next step withoutpurification.

I-4: benzyl (4-carbamoylbicyclo[2.2.1]heptan-1-yl)carbamate

To a solution of compound I-3 (2 g, 6.9 mmol) and Et₃N (1 g, 10 mmol) inDCM (20 mL) was added isobutyl chloroformate (1.36 g, 10 mmol) dropwiseat 0° C. The mixture was stirred at 0° C. for 20 min, added with NH₄OH(10 mL) dropwise and then stirred at room temperature for 10 min. Theresulting mixture was poured into water (30 mL), and the organic phasewas separated. The aqueous solution was extracted with DCM (15 mL×2),and the combined organic phases were dried over Na₂SO₄, filtered andconcentrated. The residue was purified by column chromatography (eluent:EA/PE (1:1)-EA/MeOH (10:1)), to give the desired compound I-4 (1.7 g,yield: 85%).

I-5: benzyl (4-aminobicyclo[2.2.1]heptan-1-yl)carbamate

A solution of compound I-4 (1.6 g, 5.55 mmol) and[hydroxy(tosyloxy)iodo]benzene (2.17 g, 5.55 mmol) in ACN (20 mL) washeated to reflux for 1 h. The solvent was evaporated and 1 M NaOH (12mL) was added. FA (15 mL×2) was added for extraction, and the organicphase was dried over Na₂SO₄, filtered and concentrated. The residue waspurified by column chromatography (eluent: DCM/MeOH=10:1) to give thedesired compound I-6 (920 mg, yield: 64%). LC-MS m/z=261.1 [M+1]⁺.

Synthesis of Intermediate Boronic Acid II

Synthetic Route of Intermediate II-1

4-bromo-N-(pyridin-2-yl)benzamide

To a mixture of 4-bromobenzoic acid (5 g, 24.8 mmol) and pyridin-2-amine(4.68 g, 49 mmol) in pyridine (30 mL) was added POCl₃ (11.4 g, 74 mmol)dropwise under an ice bath. The suspension was stirred at roomtemperature for 20 min. The reaction mixture was poured into water (100mL) and extracted with ethyl acetate (40 mL×3). The organic phase waswashed with saturated aqueous NaCl solution (50 mL/2), dried overanhydrous Na₂SO₄, filtered and evaporated. The residue was purified bycolumn chromatography (ethyl acetate/petroleum ether=1:9-1:1) to givethe product 4-bromo-N-pyridin-2-yl)benzamide (3.28 g, 48%). LC-MSm/z=277.0 [M+1]⁺.

N-(pyridin-2-yl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide

A mixture of 4-bromo-N-(pyridin-2-yl)benzamide (2 g, 7.22 mmol),4,4,4′,4′,5,5,5′,5′-octamethyl-2,2′-bi(1,3,2-dioxaborolane) (2.75 g,10.83 mmol), PdCl₂(dppf) (527 mg, 0.72 mmol) and KOAc (235 mg, 2.4 mmol)in toluene (30 mL) was heated to 110° C. and maintained at thistemperature for 6 h. The reaction mixture was evaporated and added withwater (100 mL). Ethyl acetate (40 mL×2) was added for extraction. Theorganic phase was separated, dried over anhydrous Na₂SO₄, filtered andevaporated. The residue was purified by column chromatography (ethylacetate/petroleum ether=1:4-1:1) to give the productN-(pyridin-2-yl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide(2 g, 85%).

II-1: (4-(pyridin-2-ylcarbamoyl)phenyl)boronic acid

To a solution ofN-(pyridin-2-yl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide (2 g, 6.2 mmol) in a mixed solvent of THF:H₂O (24 mL:6 mL) wasadded NaIO₄ (3.27 g, 18.6 mmol), and the mixture was stirred at roomtemperature for 30 min. 2 N aqueous HCl solution (1.65 mL) was thenadded, and the resulting mixture was stirred at room temperature for 3h. The mixture was diluted with ethyl acetate and washed with brine. Theorganic phase was separated, dried over anhydrous Na₂SO₄, filtered andconcentrated. The residue was purified by column chromatography(MeOH/DCM=1:10) to give the product(4-(pyridin-2-ylcarbamoyl)phenyl)boronic acid (II-1) (1.4 g, 93%). LC-MSm/z=243.1 [M+1]⁺.

Synthetic Route of Intermediate II-2

4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzoyl chloride

To a solution of 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzoicacid (10 g, 40 mmol) and 1 drop of DMF in DCM (100 mL) was added oxalylchloride (10.2 g, 80 mmol) dropwise under an ice bath. The mixture wasstirred at 0° C. for 30 min, and then warmed to room temperature andmaintained at this temperature for 3 h. The mixture was thenconcentrated to give a product, which was directly used in the next stepwithout purification.

N-(4-fluoropyridin-2-yl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide

To a solution of 4-fluoropyridin-2-amine (421 mg, 3.76 mmol) in pyridine(3 mL) was added a solution of4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-2-yl)benzoylchloride (1 g, 3.76 mmol) in DCM (6 mL), and the suspension was stirredat 0° C. for 30 min. The mixture was poured into water and extractedwith DCM (20 mL×2). The organic phase was washed with saturated aqueousNaCl solution, separated, dried over anhydrous Na₂SO₄ and evaporated,and the residue was purified by column chromatography (ethylacetate/petroleum ether=1:9) to give the product (1.04 g, 81%). LC-MSm/z=343.2 [M+1]⁺.

II-2: (4-((4-fluoropyridin-2-yl)carbamoyl)phenyl)boronic acid

To a solution ofN-(4-fluoropyridin-2-yl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide (1.04 g, 3.04 mmol) in a mixed solvent of THF:H₂O (24 mL:6 mL)was added NaIO₄ (1.9 g, 9.12 mmol), and the mixture, was stirred at roomtemperature for 30 min. Aqueous HCl solution (1.65 mL) was then added,and the resulting mixture was stirred at room temperature for 3 h. Themixture was diluted with ethyl acetate and washed with brine. Theorganic phase was separated, dried over anhydrous Na₂SO₄, filtered andconcentrated. The residue was purified by column chromatography(MeOH/DCM=1:10) to give the product II-2 (648 mg, 82%). LC-MS m/z=261.1[M+1]⁺.

II-3: (4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)boronic acid

With 4-(trifluoromethyl)pyridin-2-amine (609 mg, 3.76 mmol) and4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-2-yl)benzoylchloride (1 g, 3.76 mmol) as the starting materials, the same syntheticmethod as that of II-2 was used to give the desired compound (607 mg).LC-MS m/z=311.1 [M+1]⁺.

II-4: (4-((4-methylpyridin-2-yl)carbamoyl)phenyl)boronic acid

With 4-methyl-pyridin-2-amine (406 mg, 3.76 mmol) and4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-2-yl)benzoylchloride (1 g, 3.76 mmol) as the starting materials, the same syntheticmethod as that of II-2 was used to give the desired compound (589 mg).LC-MS m/z=257.1 [M+1]⁺.

II-5: (4-((4-cyanopyridin-2-yl)carbamoyl)phenyl)boronic acid

With 4-cyano-pyridin-2-amine (447 mg, 3.76 mmol) and4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-2-yl)benzoylchloride (1 g, 3.76 mmol) as the starting materials, the same syntheticmethod as that of II-2 was used to give the desired compound (465 mg).LC-MS m/z=268.0 [M+1]⁺.

Synthetic Route of Intermediate II-6

4-bromo-2-fluoro-N-(pyridin-2-yl)benzamide

To a solution of 4-bromo-2-fluorobenzoic acid (1 g, 4.56 mmol) in DCM(30 mL) was added oxalyl chloride (1.16 g, 9.13 mmol) dropwise under anice bath, and then 1 drop of DMF was added. After being stirred at roomtemperature for 3 h, the mixture was concentrated and dissolved in DCM(6 mL). The resulting solution was added to a solution ofpyridin-2-amine (428 mg, 4.56 mmol) in pyridine (3 mL) at 0° C., and thesuspension was stirred at 0° C. for 30 min. The mixture was poured intowater and extracted with DCM (20 mL×2). The organic phase was washedwith saturated aqueous NaCl solution, separated, dried over anhydrousNa₂SO₄ and evaporated, and the residue was purified by columnchromatography (ethyl acetate/petroleum ether=1:9) to give the product(1.05 g, 78%). LC-MS m/z=295.0 [M+1]⁺.

2-fluoro-N-(pyridin-2-yl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide

A solution of 4-bromo-2-fluoro-N-(pyridin-2-yl)benzamide (1.05 g, 3.55mmol), (4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3,2-dioxaborolane(1.36 g, 5.3 mmol). PdCl₂(dppf) (260 mg, 0.36 mmol) and KOAc (1.04 g,10.65 mmol) in toluene (30 mL) was heated to 110° C. and maintained atthis temperature for 6 h. The reaction mixture was evaporated and thenadded with water (100 mL). Ethyl acetate (40 mL×2) was added forextraction. The organic phase was separated, dried over anhydrousNa₂SO₄, filtered and evaporated. The residue was purified by columnchromatography (ethyl acetate/petroleum ether=1:4-1:1) to give theproduct (971 mg, 80%).

II-6: (3-fluoro-4-(pyridin-2-ylcarbamoyl)phenyl)boronic acid

To a solution of2-fluoro-N-(pyridin-2-yl)-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzamide(970 mg, 2.84 mmol) in a mixed solvent of THF:H₂O (24 mL:6 mL) was addedNaIO₄ (1.8 g, 8.52 mmol), and the mixture was stirred at roomtemperature for 30 min. Aqueous HCl solution (1.65 mL) was then added.After being stirred at room temperature for 3 h, the mixture was dilutedwith ethyl acetate and washed with brine. The organic phase wasseparated, dried over anhydrous Na₂SO₄, filtered and concentrated. Theresidue was purified by column chromatography (MeOH/DCM=1:10) to givethe product II-6 (605 mg, 82%). LC-MS m/z=261.1 [M+1]⁺.

II-7: (4-(thiazol-2-ylcarbamoyl)phenyl)boronic acid

With thiazol-2-amine (376 mg, 3.76 mmol) and4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-2-yl)benzoylchloride (1 g, 3.76 mmol) as the starting materials, the same syntheticmethod as that of II-2 was used to give the desired compound II-7 (580mg). LC-MS m/z=249.1 [M+1]⁺.

II-8: (2-fluoro-4-(pyridin-2-ylcarbamoyl)phenyl)boronic acid

With 4-bromo-3-fluoro-benzoic acid and pyridin-2-amine as the startingmaterials, the same synthetic method as that of II-1 was used to givethe desired compound II-8 (138 mg). LC-MS m/z=261.0 [M+1]⁺.

II-9:(2-fluoro-4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)boronicacid

With 4-bromo-3-fluoro-benzoic acid and4-(trifluoromethyl)-pyridin-2-amine as the starting materials, the samesynthetic method as that of II-1 was used to give the desired compoundII-9 (130 mg). LC-MS m/z=329.0 [M+1]⁺.

II-10:(2-methoxy-4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)boronicacid

With 4-bromo-3-methoxybenzoic acid (I g, 4.36 mmol) and4-(trifluoromethyl)-pyridin-2-amine (706 mg, 4.36 mmol) as the startingmaterials, the same synthetic method as that of II-6 was used to givethe desired compound II-10 (440 mg). LC-MS m/z=341.0 [M+1]⁺.

Synthetic Route of Intermediate Borate II-11

N-(4-bromobenzyl)-2-methoxybenzamide

A solution of 4-bromobenzylamine (1 g, 5.38 mmol), 2-methoxy benzoicacid (818 mg, 5.38 mmol), HATU (2.45 g, 6.46 mmol) and DIEA (1.39 g,10.76 mmol) in DMF (20 mL) was stirred at room temperature for 2 h. Themixture was poured into water (50 mL), filtered, washed with water (30mL×2) and dried to give the desired compound (1.6 g, yield: 93%), whichwas directly used in the next step without purification.

II-11: 2-methoxy-N-(4(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl)benzamide

A solution of N-(4-bromobenzyl)-2-methoxy benzamide (1.6 g, 5 mmol),4,4,4′,4′,5,5,5′,5′-octamethyl-2,2′-bi(1,3,2-dioxaborolane) (1.9 g, 7.5mmol), PdCl₂(dppf) (365 mg, 0.5 mmol) and KOAc (1.47 g, 15 mmol) indioxane (30 mL) was heated to 100° C. and maintained at this temperaturefor 6 h. The mixture was concentrated, added with water (100 ml), andthen extracted with ethyl acetate (20 mL×2). The organic phase wasseparated, dried over Na₂SO₄, filtered and concentrated, and the residuewas purified by column chromatography (ethyl acetate/petroleum ether1:9) to give the desired compound II-11 (1.5 g, yield: 82%).

II-12:5-fluoro-2-methoxy-N-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl)benzamide

With 5-fluoro-2-methoxybenzoic acid (915 mg, 5.38 mmol) and4-bromobenzylamine (I g, 5.38 mmol) as the starting materials, the samesynthetic method as that of II-11 was used to give the desired compoundII-12 (900 mg).

II-13:4-fluoro-2-methoxy-N-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl)benzamide

With 4-fluoro-2-methoxybenzoic acid (915 mg, 5.38 mmol) and4-bromobenzylamine (1 g, 5.38 mmol) as the starting materials, the samesynthetic method as that of II-11 was used to give the desired compoundII-13 (1 g).

Synthetic Route of Intermediate A-4

A-1:benzyl(4-(((3-chloropyrazin-2-yl)methyl)carbamoyl)bicyclo[2.2.1]heptan-1-yl)carbamate

A solution of4-(((benzyloxy)carbonyl)amino)bicyclo[2.2.1]heptane-1-carboxylic acid (2g, 6.9 mmol), HATU (2.89 g, 7.6 mmol), DIEA (3.56 g, 27.6 mmol) and thehydrochloride of (3-chloropyrazin-2-yl)methylamine (1.3 g, 7.24 mmol) inDMF (20 mL) was stirred at room temperature for 6 h. The mixture waspoured into water (100 mL) and extracted with ethyl acetate (30 mL×2).The organic phase was washed with saturated aqueous NaCl solution,separated, dried over Na₂SO₄, filtered and evaporated. The residue waspurified by column chromatography (ethyl acetate/petroleumether=1:1-1:0) to give the desired compound A-1 (2 g, yield: 70%). LC-MSm/z=414.9 [M+1]⁺.

A-2: benzyl(4-(8-chloroimidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

To a solution of benzyl(4N-(((3-chloropyrazin-2-yl)methyl)carbamoyl)bicyclo[2.2.1]heptan-1-yl)carbamate(A-1) (2 g, 4.83 mmol) in ACN (30 mL) were added pyridine (381 mg, 4.83mmol) and PCI₅ (4 g, 19.32 mmol), and the mixture was heated to 56° C.and maintained at this temperature for 1 h. After being cooled to roomtemperature, the mixture was slowly poured into ice saturated aqueousNaHCO₃ solution (100 mL). While maintaining pH at 9, the mixture wasextracted with ethyl acetate (30 mL×2). The organic phase was washedwith saturated aqueous NaCl solution, separated, dried over Na₂SO₄,filtered and evaporated. The residue was purified by columnchromatography (ethyl acetate/petroleum ether=1:1) to give the desiredcompound A-2 (1.5 g, yield: 78%).

A-3: benzyl(4-(8-chloro-1-iodoimidazo[1,5-c]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

A mixed solution of benzyl(4-(8-chloroimidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)carbamate(A-2) (1.5 g, 3.79 mmol) and NIS (1.13 g, 5.04 mmol) in DMF (10 mL) washeated to 60° C. under N₂ atmosphere and stirred for 10 h. After beingcooled to room temperature, the mixture was poured into water (100 mL)and extracted with ethyl acetate (30 mL×2). The organic phase was washedwith saturated aqueous NaCl solution, separated, dried over Na₂SO₄,filtered and evaporated. The residue was purified by columnchromatography (ethyl acetate/petroleum ether=2:3) to give the desiredcompound A-3 (1.52 g, yield: 77%).

A-4: benzyl(4-(8-amino-1-iodoimidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

To a suspension of compound A-3 (1.5 g, 2.87 mmol) in IPA (15 mL) wasadded NH₄OH (3 mL), and the mixture was heated to 110° C. and maintainedat this temperature for 6 h. The mixture was then concentrated and addedwith saturated aqueous NaHCO₃ solution (20 mL). Ethyl acetate (30 mL×2)was added for extraction. The organic phase was washed with saturatedaqueous NaCl solution, separated, dried over Na₂SO₄, filtered andevaporated. The residue was purified by column chromatography (ethylacetate/petroleum ether:=1:1) to give the desired compound A-4 (1.1 g,yield: 77%).

Synthetic Route of Intermediate B-3

B-1:benzyl(4-(4-chloro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

A solution of 2-(4,6-dichloropyrimidin-5-yl)acetaldehyde (735 mg, 3.8mmol), benzyl (4-aminobicyclo[2.2.1]heptan-1-yl)carbamate (1 g, 3.8mmol) and Et₃N (389 mg, 3.8 mmol) in EtOH (20 mL) was heated to 80° C.and maintained at this temperature for 16 h. The mixture wasconcentrated, added with water (20 mL), and extracted with ethyl acetate(20 mL×2). The organic phase was separated, dried over Na₂SO₄, filteredand evaporated, and the residue was purified by column chromatography(EA/PE=1:4) to give the desired compound B-1 (1.25 g, yield: 83%). LC-MSm/z=397.1 [M+1]⁺.

B-2: benzyl(4-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

To a solution of compound B-1 (1.25 g, 3.16 mmol) in DMF (10 mL) wasadded NIS (950 mg, 4.2 mmol), and the mixture was heated to 60° C. andmaintained at this temperature for 6 h. The mixture was poured intowater (20 mL) and extracted with ethyl acetate (20 mL×2). The organicphase was separated, dried over Na₂SO₄, filtered and evaporated, and theresidue was purified by column chromatography (EA/PE=1:4) to give thedesired compound (0.09 g, yield: 66%). LC-MS m/z=523.1 [M+1]⁺.

B-3: benzyl(4-(4-amino-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

To a solution of benzyl(4-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.1]heptan-1-yl)carbamate(B-2) (1.09 g, 2.08 mmol) in IPA (10 mL) was added NH₄OH (2 mL), and themixture was heated to 110° C. and maintained at this temperature for 6h. The mixture was then concentrated, poured into aqueous NaHCO₃solution, and extracted with DCM (20 mL×2). The organic phase wasseparated, dried, filtered and concentrated, and the residue waspurified by column chromatography (MeOH/DCM=1:20) to give the desiredcompound B-3 (900 mg, yield: 86%).

Synthetic Route of Intermediate C-4

C-1: benzyl(4-((6-chloro-5-nitropyrimidin-4-yl)amino)bicyclo[2.2.1]heptan-1-yl)carbamate

A solution of 4,6-dichloro-5-nitropyrimidine (518 mg, 2.68 mmol), benzyl(4-aminobicyclo[2.2.1]heptan-1-yl)carbamate (1-5) (698 mg, 2.68 mmol)and Et₃N (1.1 g, 10.72 mmol) in DCM (20 mL) was stirred at roomtemperature for 4 h. The solvent was removed, and the residue wastreated with ethyl acetate (50 mL) and washed with saturated aqueousNaCl solution. The organic phase was separated, dried over Na₂SO₄,filtered and concentrated, and the residue was purified by columnchromatography (EA/PE=1:4) to give the desired compound C-1 (633 mg,yield: 57%).

C-2: benzyl(4-((5-amino-6-chloropyrimidin-4-yl)amino)bicyclo[2.2.1]heptan-1-yl)carbamate

To a solution of compound C-1 (400 mg, 0.96 mmol) in a mixed solvent(EtOH/H₂O=20 mL/4 mL) were added Fe powder (268 mg, 4.8 mmol) and NH₄Cl(254 mg, 4.8 mmol) The mixture was then heated to reflux for 1 h. Afterbeing cooled to room temperature, the mixture was filtered and washedwith MeOH (10 mL). The filtrate was concentrated, and the crude productwas purified by column chromatography (EA/PE=1:4) to give the desiredcompound C-2 (292 mg, yield: 79%).

C-3: benzyl(4-(6-chloro-8-oxo-7,8-dihydropurin-9-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

To a solution of compound C-2 (290 mg, 0.75 mmol) in DCM (10 mL) at 0°C. were added Et₃N (166 mg, 1.5 mmol) and triphosgene (291 mg, 0.9mmol), and the mixture was stirred at 0° C. for 2 h. The mixture wasthen poured into water (20 mL) and the organic phase was separated, andthen the aqueous solution was extracted with DCM (10 mL×2) to separatethe organic phase. The combined organic phases were dried over Na₂SO₄,filtered and concentrated to give crude C-3 (316 mg), which was directlyused in the next step without purification.

C-4: benzyl(4-(6-amino-8-oxo-7,8-dihydropurin-9-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

To a solution of compound C-3 (310 mg, 0.75 mmol) in IPA (10 mL) wasadded NH₄OH (2 mL), and the mixture was heated to 150° C. and maintainedat this temperature for 24 h. The mixture was then concentrated, pouredinto aqueous NaHCO₃ solution and extracted with DCM (20 mL×2). Theorganic phase was separated, dried, filtered and concentrated, and theresidue was purified by column chromatography (MeOH/DCM=1:20) to givethe desired compound C-4 (100 mg, yield: 34%). LC-MS m/z=395.1 [M+1]⁺.

Synthetic Route of Intermediate D-5

D-1: benzyl(4-((6-cloro-5-formylpyrimidin-4-yl)amino)bicyclo[2.2.1]heptan-1-yl)carbamate

A mixed solution of 1-5 (1.47 g, 5.65 mmol),4,6-dichloropyrimidine-5-carbaldehyde (I g, 5.65 mmol) and triethylamine(1.15 g, 11.4 mmol) in DCM (20 ml) was stirred overnight at roomtemperature. The reaction mixture was diluted with EA (50 mL) and washedwith water (30 ml×2). The organic phase was dried over anhydrous Na₂SO₄,filtered and concentrated. The residue was purified by silica gel columnchromatography (eluent: petroleum ether/ethyl acetate=1:1) to give thedesired product D-1 (1.24 g, 55%). LC-MS m/z=401.0 [M+1]⁺.

D-2: benzyl(E)(4-((6-chloro-5-((hydroxyimino)methyl)pyrimidin-4-yl)amino)bicyclo[2.2.1]heptan-1-yl)carbamate

A mixed solution of D-1 (1.2 g, 3.0 mmol), hydroxylamine-O-sulfonic acid(0.41 g, 3.6 mmol) in DCM/ACN (50 mL/50 mL) was stirred at roomtemperature for 16 h and then stirred at 50° C. for 6 h. After beingcooled, the mixture was concentrated to 10 mL, and the concentrate wasfiltered and washed with ACN (2 mL) to give the desired product D-2 (1.0g, 81%). LC-MS m/z=416.0 [M+1]⁺.

D-3: benzyl(4-(4-chloro-1H-pyrazolo[3,4-d]pyrimidin-1-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

To a solution of D-2 (1.0 g, 2.41 mmol) in DCM (100 mL) was added DIEA(4 mL), and then TsCl (0.23 mL, 2.9 mmol) was added dropwise. Afterbeing stirred at room temperature for 3 h, the reaction mixture wastreated with water (100 mL). The organic phase was washed with brine (30mL×2), dried over anhydrous Na₂SO₄, filtered and concentrated. Theresidue was purified by silica gel column chromatography (eluent: ethylacetate/petroleum ether=1:6) to give the desired product D-3 (400 mg,42%). LC-MS m/z=398.1 [M+1]⁺.

D-4: benzyl(4-(4-amino-1H-pyrazolo[3,4-d]pyrimidin-1-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

A mixture of D-3 (4 (0) mg, 1.0 mmol) and ammonium hydroxide (30%, 5 mL)in isopropanol (20 mL) was stirred in a sealed tube at 120° C. for 6 h.The solvent was evaporated off and the residue was purified by silicagel column chromatography (eluent: PE/EA=2:1) to give the desiredproduct D-4 (280 mg, 73.4%).

D-5: benzyl(4-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

A mixture of D-4 (190 mg, 0.5 mmol), NIS (4 (0) mg, 1.78 mmol) and HBF₄(50%, 19.6 mmol, 4 mL) in ACN (2.5 mL) was heated to 85° C. in a sealedtube and stirred for 6 h. After the mixture was cooled, the reaction wasquenched with saturated NaHCO₃ and EA (50 mL×2) was added forextraction. The organic phase was dried over anhydrous Na₂SO₄, filteredand concentrated. The residue was purified by silica gel columnchromatography (eluent: PE/EA=1: 1-1:0) to give the desired product D-5(96 mg, 38.1%). LC-MS m/z=505.0 [M+1]⁺.

Example 1 Synthetic Route of Compound A-7-n

A-5:3-(4-aminobicyclo[2.2.1]heptan-1-yl)-1-iodoimidazo[1,5-a]pyrazin-8-amine

A solution of compound A-4 (1 g, 1.99 mmol) in a mixed solvent ofTFA/DCM (10 mL/10 mL) was heated to 60° C. and maintained at thistemperature for 6 h. The mixture was concentrated and added with DCM (20mL×2). The resulting mixture was concentrated, dissolved in DCM (20 mL),added with a solution of HCl in dioxane (5 mL), and stirred at roomtemperature for 10 min. The mixture was evaporated and added with DCM(20 mL×2), and the resulting mixture was evaporated and added with DME(20 mL). The mixture was stirred at room temperature for 30 min.filtered and then washed with DME (10 mL×2). The obtained compound A-5was directly used in the next step without purification. LC-MS m/z=370.1[M+1]⁺.

A-6:N-(4-(8-amino-1-iodoimidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)but-2-ynamide

A solution of compound A-S (1.35 g, 2.8 mmol). DIEA (3.28 g, 25.2 mmol),but-2-ynoic acid (235 mg, 2.8 mmol) and HATU (1.06 g, 2.8 mmol) in DMF(20 mL) was stirred at room temperature for 30 min. The mixture waspoured into water (30 mL) and extracted with ethyl acetate (30 mL×2).The organic phase was washed with saturated aqueous NaCl solution,separated, dried over Na₂SO₄, filtered and evaporated. The residue waspurified by column chromatography (ethyl acetate/petroleum ether:=2:3)to give the desired compound A-6 (800 mg, yield: 65%).

A-7-1:4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(pyridin-2-yl)benzamide

A solution of compound A-6 (30 mg, 0.069 mmol),(4-(pyridin-2-ylcarbamoyl)phenyl) boronic acid (II-1) (20 mg, 0.085mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃ (45 mg, 0.138 mmol) in amixed solvent of DME/H₂O (1.5 mL/0.3 mL) was heated to 80° C. andmaintained at this temperature for 3 h. The mixture was concentrated,added with water (20 mL) and extracted with ethyl acetate (20 mL×2). Theorganic phase was separated, dried over Na₂SO₄, filtered and evaporated.The residue was purified by column chromatography (ethylacetate/petroleum ether=1:1) to give the desired compound A-7-1 (10 mg,yield: 30%). LC-MS m/z=506.2 [M+1]⁺.

A-7-2:4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-fluoropyridin-2-yl)benzamide

A solution of compound A-6 (30 mg, 0.069 mmol),(4-((4-fluoropyridin-2-yl)carbamoyl) phenyl)boronic acid (II-2) (22 mg,0.085 mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃ (45 mg, 0.138mmol) in a mixed solvent of DMF/H₂O (1.5 mL/0.3 mL) was heated to 80° C.and maintained at this temperature for 3 h. The mixture wasconcentrated, added with water (20 mL) and extracted with ethyl acetate(20 mL×2). The organic phase was separated, dried over Na₂SO₄, filteredand evaporated. The residue was purified by column chromatography (ethylacetate/petroleum ether=1:1) to give the desired compound A-7-2 (15 mg,yield: 34%). LC-MS m/z=524.0 [M+1]⁺.

A-7-3:4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound A-6 (30 mg, 0.069 mmol),(4-((4-(trifluoromethyl)pyridin-2-yl) carbamoyl)phenyl)boronic acid(II-3) (26 mg, 0.085 mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃ (45mg, 0.138 mmol) in a mixed solvent of DME/H₂O (1.5 mL/0.3 mL) was heatedto 140° C. and maintained at this temperature for 3 h. The mixture wasconcentrated, added with water (20 mL) and extracted with ethyl acetate(20 mL×2). The organic phase was separated, dried over Na₂SO₄, filteredand evaporated. The residue was purified by column chromatography (ethylacetate/petroleum ether=1:1) to give the desired compound A-7-3 (12 mg,yield: 31%). LC-MS m/z=574.2 [M+1]⁺.

A-7-4:4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-methylpyridin-2-yl)benzamide

A solution of compound A-6 (30 mg, 0.069 mmol),(4-((4-methylpyridin-2-yl)carbamoyl) phenyl)boronic acid (114) (22 mg,0.085 mmol). Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃ ₍45 mg, 0.138mmol) in a mixed solvent of DMF/H₂O (1.5 mL/0.3 mL) was heated to 80° C.and maintained at this temperature for 3 h. The mixture wasconcentrated, added with water (20 mL) and extracted with ethyl acetate(20 mL×2). The organic phase was separated, dried over Na₂SO₄, filteredand evaporated. The residue was purified by column chromatography (ethylacetate/petroleum ether=1:1) to give the desired compound A-7-4 (14 mg,yield: 39%). LC-MS m/z=520.2 [M+1]⁺.

A-7-5:4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-cyanopyridin-2-yl)benzamide

A solution of compound A-6 (30 mg, 0.069 mmol),(4-((4-cyanopyridin-2-yl)carbamoyl) phenyl)boronic acid (II-5) (23 mg,0.085 mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃ (45 mg, 0.138mmol) in a mixed solvent of DME/H₂O (1.5 mL/0.3 mL) was heated to 80° C.and maintained at this temperature for 3 h. The mixture wasconcentrated, added with water (20 mL) and extracted with ethyl acetate(20 mL 2). The organic phase was separated, dried over Na₂SO₄, filteredand evaporated. The residue was purified by column chromatography (ethylacetate/petroleum ether=1:1) to give the desired compound A-7-5 (12 mg,yield: 33%). LC-MS m/z=531.0 [M+1]⁺.

A-7-6: 4-(8-amino-3(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-2-fluoro-N-(pyridin-2-yl)benzamide

A solution of compound A-6 (30 mg, 0.069 mmol),(3-fluoro-4-(pyridin-2-ylcarbamoyl) phenyl)boronic acid (II-6)(22 mg,0.085 mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃ (45 mg, 0.138mmol) in a mixed solvent of DME/H₂O (1.5 mL/0.3 mL) was heated to 80° C.and maintained at this temperature for 3 h. The mixture wasconcentrated, added with water (20 mL) and extracted with ethyl acetate(20 mL×2). The organic phase was separated, dried over Na₂SO₄, filteredand evaporated. The residue was purified by column chromatography (ethylacetate/petroleum ether=1:1) to give the desired compound A-7-6 (15 mg,yield: 42%). LC-MS m/z=524.2 [M+1]⁻.

A-7-7:4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(thiazol-2-yl)benzamide

A solution of compound A-6 (30 mg, 0.069 mmol),(4-(thiazol-2-ylcarbamoyl)phenyl) boronic acid (II-7)(28 mg, 0.085mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃ (45 mg, 0.138 mmol) in amixed solvent of DME/H₂O (1.5 mL/0.3 mL) was heated to 80° C. andmaintained at this temperature for 3 h. The mixture was concentrated,added with water (20 mL) and extracted with ethyl acetate (20 mL×2). Theorganic phase was separated, dried over Na₂SO₄, filtered and evaporatedto dryness. The residue was purified by column chromatography (ethylacetate/petroleum ether=1:1) to give the desired compound A-7-7 (13 mg,yield: 37%). LC-MS m/z=512.0 [M+1]⁺.

A-7-8:4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-3-fluoro-N-(pyridin-2-yl)benzamide

A solution of compound A-6 (30 mg, 0.069 mmol),(2-fluoro-4-(pyridin-2-ylcarbamoyl) phenyl)boronic acid (II-8) (22 mg,0.085 mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃ (45 mg, 0.138mmol) in a mixed solvent of DME/H₂O (1.5 mL/0.3 mL) was heated to 80° C.and maintained at this temperature for 3 h. The mixture wasconcentrated, added with water (20 mL) and extracted with ethyl acetate(20 mL×2). The organic phase was separated, dried over Na₂SO₄, filteredand evaporated. The residue was purified by column chromatography (ethylacetate/petroleum ether=1:1) to give the desired compound A-7-8 (14 mg,yield: 39%). LC-MS m/z=524.0 [M+1]⁺.

A-7-9:4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-3-fluoro-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound A-6 (30 mg, 0.069 mmol),(2-fluoro-4-((4-(trifluoromethyl) pyridin-2-yl)carbamoyl)phenyl)boronicacid (II-9) (28 mg, 0.085 mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) andCs₂CO₃ (45 mg, 0.138 mmol) in a mixed solvent of DME/H₂O (1.5 mL/0.3 mL)was heated to 80° C. and maintained at this temperature for 3 h. Themixture was concentrated, added with water (20 mL) and extracted withethyl acetate (20 mL×2). The organic phase was separated, dried overNa₂SO₄, filtered and evaporated. The residue was purified by columnchromatography (ethyl acetate/petroleum ether=1:1) to give the desiredcompound A-7-9 (19 mg, yield: 47%). LC-MS m/z=592.0 [M+1]⁺.

A-7-10:4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-3-methoxy-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound A-6 (30 mg, 0.069 mmol),(2-methoxy-4-((4-(trifluoromethyl) pyridin-2-yl)carbamoyl)phenyl)boronicacid (II-10) (29 mg, 0.085 mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) andCs₂CO₃ (45 mg, 0.138 mmol) in a mixed solvent of DME/H₂O (1.5 mL/0.3 mL)was heated to 80° C. and maintained at this temperature for 3 h. Themixture was concentrated, added with water (20 mL) and extracted withethyl acetate (20 mL×2). The organic phase was separated, dried overNa₂SO₄, filtered and evaporated. The residue was purified by columnchromatography (ethyl acetate/petroleum ether=1:1) to give the desiredcompound A-7-10 (16 mg, yield: 38%). LC-MS m/z=604.0 [M+1]⁺.

A-7-11:N-(4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)benzyl)-2-methoxybenzamide

A solution of compound A-6 (30 mg, 0.069 mmol),2-methoxy-N-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl)benzamide(II-11) (31 mg, 0.085 mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃(45 mg, 0.138 mmol) in a mixed solvent of DME/HO (1.5 mL/0.3 mL) washeated to 80° C. and maintained at this temperature for 3 h. The mixturewas concentrated, added with water (20 mL) and extracted with ethylacetate (20 mL×2). The organic phase was separated, dried over Na₂SO₄,filtered and evaporated. The residue was purified by columnchromatography (ethyl acetate/petroleum ether:=1:1) to give the desiredcompound A-7-11 (21 mg, yield: 50%). LC-MS m/z=549.3 [M+1]⁺.

A-7-12:N-(4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)benzyl)-5-fluoro-2-methoxybenzamide

A solution of compound A-6 (30 mg, 0.069 mmol),5-fluoro-2-methoxy-N-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl)benzamide(II-12) (33 mg, 0.085 mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃(45 mg, 0.138 mmol) in a mixed solvent of DME/H₂O (1.5 mL/0.3 mL) washeated to 80° C. and maintained at this temperature for 3 h. The mixturewas concentrated, added with water (20 mL) and extracted with ethylacetate (20 mL×2). The organic phase was separated, dried over anhydrousNa₂SO₄, filtered and evaporated. The residue was purified by columnchromatography (ethyl acetate/petroleum ether=1:1) to give the desiredcompound A-7-12 (24 mg, yield: 56%). LC-MS m/z=567.0 [M+1]⁺.

A-7-13:N-(4-(8-amino-3-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)benzyl)-4-fluoro-2-methoxybenzamide

A solution of compound A-6 (30 mg, 0.069 mmol),4-fluoro-2-methoxy-N-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzil)benzamide(II-13) (33 mg, 0.085 mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃(45 mg, 0.138 mmol) in a mixed solvent of DME/H₂O (1.5 mL/0.3 mL) washeated to 80° C. and maintained at this temperature for 3 h. The mixturewas concentrated, added with water (20 mL) and extracted with ethylacetate (20 mL×2). The organic phase was separated, dried over Na₂SO₄,filtered and evaporated. The residue was purified by columnchromatography (ethyl acetate/petroleum ether=1:1) to give the desiredcompound A-7-13 (26 mg, yield: 60%). LC-MS m/z=567.1 [M+1]+.

A-7-14:N-(4-(8-amino-1-(4-phenoxyphenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)but-2-ynamide

A solution of compound A-6 (30 mg, 0.069 mmol), 4-phenoxyphenylboronicacid (18 mg, 0.085 mmol), Pd[PPh₃]₄ (8 mg, 0.0069 mmol) and Cs₂CO₃ (45mg, 0.138 mmol) in a mixed solvent of DME/H₂O (1.5 m/0.3 mL) was heatedto 80° C. and maintained at this temperature for 3 h. The mixture wasconcentrated, added with water (20 mL) and extracted with ethyl acetate(20 mL×2). The organic phase was separated, dried over Na₂SO₄, filteredand evaporated to dryness. The residue was purified by columnchromatography (ethyl acetate/petroleum ether=1:1) to give the desiredcompound A-7-14 (19 mg, yield: 49%). LC-MS m/z=478.0 [M+1]+.

Synthetic Route of Compound A-10-n

A-8: benzyl(4-(8-amino-1-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

A solution of benzyl(4-(8-amino-1-iodoimidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)carbamate(A-4) (300 mg, 0.6 mmol), (4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)boronic acid (II-3) (229 mg, 0.738 mmol), Pd[PPh₃]₄ (69mg, 0.06 mmol) and Cs₂O₃ (239 mg, 0.738 mmol) in a mixed solvent ofDME:H₂O (2.5 mL:0.5 mL) was heated to 80° C. and stirred overnight. Themixture was concentrated, and the residue was purified by columnchromatography (methanol/DCM=1:30) to give the desired compound A-8 (265mg, yield: 69%).

A-9: 4(8-amino-3-(4-aminobicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound A-8 (265 mg, 0.42 mmol) in a mixed solvent ofDCM/TFA (10 mL:10 mL) was heated to 60° C. and stirred for 18 h. Afterthe mixture was evaporated to dryness, DCM (20 mL×2) was added, and theresulting mixture was concentrated. The residue was dissolved in DCM (30mL), and then the solution was added to a solution of HCl in dioxane.After the mixture was evaporated to dryness, DCM (20 mL×2) was thenadded, and the resulting mixture was concentrated, followed by theaddition of isopropyl ether (30 mL). After being stirred for 2 h, themixture was filtered and washed with isopropyl ether (10 mL×2) to givethe hydrochloride of the desired product A-9 (19 mg), which was directlyused in the next step without purification.

A-10-1:(l)-4-(8-amino-3-(4-(4-methoxybut-2-enamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound A-9 (15 mg, 0.024 mmol). HATU (9.12 mg, 0.024mmol), DIEA (16 mg, 0.12 mmol) and (E)-4-methoxybut-2-enoic acid (2.8mg, 0.024 mmol) in DMF (1 mL) was stirred at room temperature for 1 h,and then the mixture was poured into water (5 mL) and extracted withethyl acetate (5 mL×2). The organic phase was washed with saturatedaqueous NaCl solution, separated, dried over anhydrous Na₂SO₄, filteredand evaporated to dryness. The residue was purified by columnchromatography (eluent: DCM/MeOH=20:1) to give the product A-10-1 (5 mg,yield: 35%). LC-MS m/z=606.1 [M+1]+.

A-10-2:(E)-4-(8-amino-3-(4-(4-(tetrahydropyrrol-1-yl)but-2-enamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024mmol), DIEA (16 mg, 0.12 mmol) and(F)-4-(tetrahydropyrrol-1-yl)-but-2-enoic acid (4 mg, 0.024 mmol) in DMF(1 ml) was stirred at room temperature for 1 h, and then the mixture waspoured into water (5 mL) and extracted with ethyl acetate (5 ml, 2). Theorganic phase was washed with saturated aqueous NaCl solution,separated, dried over anhydrous Na₂SO₄, filtered and evaporated todryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product A-10-2 (6 mg, yield: 38%). LC-MSm/z=645.0 [M+1]+.

A-10-3:4-(3-(4-acrylamidobicyclo[2.2.1]heptan-1-yl)-8-aminoimidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

Compound A-9 (15 mg, 0.024 mmol). HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and acrylic acid (2 mg, 0.024 mmol) were stirred at roomtemperature for 1 h, and then the mixture was poured into water (1 ml)and extracted with ethyl acetate (5 mL×2). The organic phase was washedwith saturated aqueous NaCl solution, separated, dried over anhydrousNa₂SO₄, filtered and evaporated to dryness. The residue was purified bycolumn chromatography (DCM/MeOH=20:1) to give the product A-10-3 (5 mg,yield: 38%). LC-MS m/z=562.0 [M+1]+.

A-10-4:4-(3-(4-acetamidobicyclo[2.2.1]heptan-1-yl)-8-aminoimidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

Compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and acetic acid (1.5 mg, 0.024 mmol) were stirred at roomtemperature for 1 h, and then the mixture was poured into water (1 mL)and extracted with ethyl acetate (5 mL×2). The organic phase was washedwith saturated aqueous NaCl solution, separated, dried over anhydrousNa₂SO₄, filtered and evaporated to dryness. The residue was purified bycolumn chromatography (DCM/MeOH=20:1) to give the product A-10-4 (4 mg,yield: 31%). LC-MS m/z=550.0 [M+1]+.

A-10-5:N-(4-(8-amino-1-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)-3-methyloxetane-3-carboxamide

A solution of compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024mmol), DIEA (16 mg, 0.12 mmol) and 3-methyloxetane-3-carboxylic acid (3mg, 0.024 mmol) in DMF (1 mL) was stirred at room temperature for 1 h,and then the mixture was poured into water (5 mL) and washed with ethylacetate (5 mL×2). The organic phase was washed with saturated aqueousNaCl solution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product A-10-5 (6 mg, yield: 40%). LC-MSm/z=606.1 [M+1]+.

A-10.6:4-(8-amino-3-(4-(2-hydroxy-2-methylpropanamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

Compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and 2-hydroxy-2-methylpropionic acid (2.5 mg, 0.024 mmol)were stirred at room temperature for 1 h, and then the mixture waspoured into water (1 mL) and extracted with ethyl acetate (5 mL×2). Theorganic phase was washed with saturated aqueous NaCl solution,separated, dried over anhydrous Na₂SO₄, filtered and evaporated todryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product A-10-6 (6 mg, yield: 41%). LC-MSm/z=594.1 [M+1]+.

A-10-7:4-(8-amino-3-(4-(2-methoxyacetamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

Compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and 2-methoxyacetic acid (2 mg, 0.024 mmol) were stirredat room temperature for 1 h, and then the mixture was poured into water(1 mL) and extracted with ethyl acetate (5 mL×2). The organic phase waswashed with saturated aqueous NaCl solution, separated, dried overanhydrous Na₂SO₄, filtered and evaporated to dryness. The residue waspurified by column chromatography (DCM/MeOH=20:1) to give the productA-10-7 (5 mg, yield: 36%). LC-MS m/z=580.1 [M+1]+.

A-10-8:4-(8-amino-3-(4-(3-methoxypropanamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

Compound A-9 (15 mg, 0.024 mmol). HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and 3-methoxypropionic acid (2.5 mg, 0.024 mmol) werestirred at room temperature for 1 h, and then the mixture was pouredinto water (1 mL) and extracted with ethyl acetate (5 mL×2). The organicphase was washed with saturated aqueous NaCl solution, separated, driedover anhydrous Na₂SO₄, filtered and evaporated to dryness. The residuewas purified by column chromatography (DCM/MeOH=20:1) to give theproduct A-10-8 (5 mg, yield: 35%). LC-MS m/z=594.1 [M+1]+.

A-10-9:4-(8-amino-3-(4-(1-hydroxycyclopropanecarboxamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024mmol), DIEA (16 mg, 0.12 mmol) and 1-hydroxycyclopropanecarboxylic acid(2.5 mg, 0.024 mmol) in DMF (1 mL) was stirred at room temperature for 1h, and then the mixture was poured into water (5 mL) and extracted withethyl acetate (5 mL×2). The organic phase was washed with saturatedaqueous NaCl solution, separated, dried over anhydrous Na₂SO₄, filteredand evaporated to dryness. The residue was purified by columnchromatography (DCM/MeOH=20:1) to give the product A-10-9 (5 mg, yield:36%). LC-MS m/z=592.01 [M+1]+.

A-10-10:4-(8-amino-3-(4-(2-morpholinoacetamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

Compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and 2-morpholinoacetic acid (3.5 mg, 0.024 mmol) werestirred at room temperature for 1 h, and then the mixture was pouredinto water (1 mL) and extracted with ethyl acetate (5 mL×2). The organicphase was washed with saturated aqueous NaCl solution, separated, driedover anhydrous Na₂SO₄, filtered and evaporated to dryness. The residuewas purified by column chromatography (DCM/MeOH=20:1) to give theproduct A-10-10 (7 mg, yield: 47%). LC-MS m/z=635.0 [M+1]+.

A-10-11:N-(4-(8-amino-1-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)tetrahydrofuran-2-carboxamide

A solution of compound A-9 (15 mg, 0.024 mmol). HATU (9.12 mg, 0.024mmol), DIEA (16 mg, 0.12 mmol) and tetrahydrofuran-2-carboxylic acid(2.8 mg, 0.024 mmol) in DMF (1 mL) was stirred at room temperature for 1h, and then the mixture was poured into water (5 mL) and extracted withethyl acetate (5 mL×2). The organic phase was washed with saturatedaqueous NaCl solution, separated, dried over anhydrous Na₂SO₄, filteredand evaporated to dryness. The residue was purified by columnchromatography (DCM/MeOH=20:1) to give the product A-10-11 (5 mg, yield:35%). LC-MS m/z=606.0 [M+1]+.

A-10-12:4-(8-amino-3-(4-(1-cyanocyclopropanecarboxamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

Compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and 1-cyanocyclopropanecarboxylic acid (2.7 mg, 0.024mmol) were stirred at room temperature for 1 h, and then the mixture waspoured into water (1 mL) and extracted with ethyl acetate (5 mL×2). Theorganic phase was washed with saturated aqueous NaCl solution,separated, dried over anhydrous Na₂SO₄, filtered and evaporated todryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product A-10-12 (6 mg, yield: 42%). LC-MSm/z=601.3 [M+1]+.

A-10-13:4-(8-amino-3-(4-(2-cyanoacetamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

Compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and 2-cyanoacetic acid (2 mg, 0.024 mmol) were stirred atroom temperature for 1 h, and then the mixture was poured into water (1mL) and extracted with ethyl acetate (5 mL×2). The organic phase waswashed with saturated aqueous NaCl solution, separated, dried overanhydrous Na₂SO₄, filtered and evaporated to dryness. The residue waspurified by column chromatography (DCM/MeOH=20:1) to give the productA-10-13 (6 mg, yield: 44%). LC-MS m/z=575.2 [M+1]+.

A-10-14:4-(8-amino-3-(4-(2-cyanopropanamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

Compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and 2-cyanopropionic acid (2.4 mg, 0.024 mmol) werestirred at room temperature for 1 h, and then the mixture was pouredinto water (1 mL) and extracted with ethyl acetate (5 mL×2). The organicphase was washed with saturated aqueous NaCl solution, separated, driedover anhydrous Na₂SO₄, filtered and evaporated to dryness. The residuewas purified by column chromatography (DCM/MeOH=20:1) to give theproduct A-10-14 (5 mg, yield: 36%). LC-MS m/z=589.3 [M+1]+.

A-10-15: N-(4-(8-amino-1-(44(4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)-4-cyanotetrahydro-2H-pyran-4-carboxamide

Compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and 4-cyanotetrahydro-2H-pyran-4-carboxylic acid (3.7 mg,0.024 mmol) were stirred at room temperature for 1 h, and then themixture was poured into water (1 mL) and extracted with ethyl acetate (5mL×2). The organic phase was washed with saturated aqueous NaClsolution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product A-10-15 (6 mg, yield: 39%). LC-MSm/z=645.0 [M+1]+.

A-10-16:1-((4-(8-amino-1-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)carbamoyl)cyclopropane-1-carboxylicacid

A solution of compound A-9 (15 mg, 0.024 mmol). HATU (9.12 mg, 0.024mmol), DIEA (16 mg, 0.12 mmol) and cyclopropane-1,1-dicarboxylic acid (3mg, 0.024 mmol) in DMF (1 mL) was stirred at room temperature for 1 h,and then the mixture was poured into water (5 mL) and extracted withethyl acetate (5 mL×2). The organic phase was washed with saturatedaqueous NaCl solution, separated, dried over anhydrous Na₂SO₄, filteredand evaporated to dryness. The residue was purified by columnchromatography (DCM/MeOH=20:1) to give the product A-10-16 (6 mg, yield:40%). LC-MS m/z=555.3 [M+1]+.

A-10-17:4-(8-amino-3-(4-benzamidobicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

Compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and benzoic acid (3 mg, 0.024 mmol) were stirred at roomtemperature for 1 h, and then the mixture was poured into water (1 mL)and extracted with ethyl acetate (5 mL×2). The organic phase was washedwith saturated aqueous NaCl solution, separated, dried over anhydrousNa₂SO₄, filtered and evaporated to dryness. The residue was purified bycolumn chromatography (DCM/MeOH=20:1) to give the product A-10-17 (8 mg,yield: 53%). LC-MS m/z=612.2 [M+1]+.

A-10-18: N-(4-(8-amino-1(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)picolinamide

Compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and 2-picolinic acid (3 mg, 0.024 mmol) were stirred atroom temperature for 1 h, and then the mixture was poured into water (1mL) and extracted with ethyl acetate (5 mL×2). The organic phase waswashed with saturated aqueous NaCl solution, separated, dried overanhydrous Na₂SO₄, filtered and evaporated to dryness. The residue waspurified by column chromatography (DCM/MeOH=20:1) to give the productA-10-18 (9 mg, yield: 60%). LC-MS m/z=613.2 [M+1]+.

A-10-19:N-(4-(8-amino-1-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)nicotinamide

Compound A-9 (15 mg, 0.024 mmol). HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and nicotinic acid (3 mg, 0.024 mmol) were stirred atroom temperature for 1 h, and then the mixture was poured into water (1mL) and extracted with ethyl acetate (5 mL×2). The organic phase waswashed with saturated aqueous NaCl solution, separated, dried overanhydrous Na₂SO₄, filtered and evaporated to dryness. The residue waspurified by column chromatography (DCM/MeOH=20:1) to give the productA-10-19 (8 mg, yield: 54%). LC-MS m/z=613.2 [M+1]+.

A-10-20:N-(4-(8-amino-1-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)-2-methoxybenzamide

Compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024 mmol), DIEA (16mg, 0.12 mmol) and 2-methoxybenzoic acid (3.6 mg, 0.024 mmol) werestirred at room temperature for 1 h, and then the mixture was pouredinto water (1 mL) and extracted with ethyl acetate (5 mL×2). The organicphase was washed with saturated aqueous NaCl solution, separated, driedover anhydrous Na₂SO₄, filtered and evaporated to dryness. The residuewas purified by column chromatography (DCM/MeOH=20:1) to give theproduct A-10-20 (8 mg, yield: 52%). LC-MS m/z=642.3 [M+1]+.

A-10-21:N-(4-(8-amino-1-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)furan-2-carboxamide

A solution of compound A-9 (15 mg, 0.024 mmol), HATU (9.12 mg, 0.024mmol), DIEA (16 mg, 0.12 mmol) and furan-2-carboxylic acid (2.7 mg,0.024 mmol) in DMF (I mL) was stirred at room temperature for 1 h, andthen the mixture was poured into water (5 mL) and extracted with ethylacetate (5 mL×2). The organic phase was washed with saturated aqueousNaCl solution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give product A-10-21 (6 mg, yield: 43%). LC-MS m/z602.2 [M+1]+.

A-10-22:N-(4-(8-amino-1-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)thiazole-2-carboxamide

A solution of compound A-9 (15 mg, 0.024 mmol). HATU (9.12 Mg, 0.024mmol), DIEA (16 mg, 0.12 mmol) and thiazole-2-carboxylic acid (3 mg,0.024 mmol) in DMF (1 mL) was stirred at room temperature for 1 h, andthen the mixture was poured into water (5 mL) and extracted with ethylacetate (5 mL×2). The organic phase was washed with saturated aqueousNaCl solution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product A-10-22 (6 mg, yield: 40%). LC-MSm/z=619.2 [M++1]+.

Synthetic Route of Compound A-13-n

A-11: benzyl(4-(8-amino-1-(4-phenoxyphenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

A solution of benzyl(4-(8-amino-1-iodoimidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)carbamate(A-4) (300 mg, 0.6 mmol), 4-phenoxybenzoic acid (158 mg, 0.738 mmol),Pd[PPh₃]₄ (69 mg, 0.06 mmol) and Cs₂O₃ (239 mg, 0.738 mmol) in a mixedsolvent of DME:H₂O (2.5 mL:0.5 mL) was heated to 80° C. and stirredovernight. The mixture was then concentrated, and the residue waspurified by column chromatography to give the desired compound A-1l (268mg, yield: 82%).

A-12:3-(4-aminobicyclo[2.2.1]heptan-1-yl)-1-(4-phenoxyphenyl)imidazo[1,5-a]pyrazin-8-amine

A solution of compound A-11 (260 mg, 0.48 mmol) in a mixed solvent ofDCM/TFA (10 mL:10 mL) was heated to 60° C. and stirred for 18 h. Afterthe mixture was evaporated to dryness, DCM (20 mL×2) was added, and theresulting mixture was concentrated. The residue was dissolved in DCM (30mL), and then the solution was added to a solution of HCl in dioxane.After the mixture was evaporated to dryness, DCM (20 mL×2) was thenadded, and the resulting mixture was concentrated, followed by theaddition of isopropyl ether (30 mL). After being stirred for 2 h, themixture was filtered and washed with isopropyl ether (10 mL×2) to givethe hydrochloride of the desired product A-12 (200 mg), which wasdirectly used in the next step without purification.

A-13-1:N-(4-(8-amino-1-(4-phenoxyphenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)-2-hydroxy-2-methylpropanamide

Compound A-12 (15 mg, 0.025 mmol), HATU (9.3 mg, 0.025 mmol), DIEA (19mg, 0.15 mmol) and 2-hydroxy-2-propionic acid (3 mg, 0.029 mmol) werestirred at room temperature for 1 h, and then the mixture was pouredinto water (1 mL) and extracted with ethyl acetate (5 mL×2). The organicphase was washed with saturated aqueous NaCl solution, separated, driedover anhydrous Na₂SO₄, filtered and evaporated to dryness. The residuewas purified by column chromatography (DCM/MeOH=20:1) to give theproduct A-13-1 (8 mg, yield: 57%). LC-MS m/z=498.4 [M+1]+.

A-13-2:N-(4-(8-amino-1-(4-phenoxyphenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)-3-methoxypropanamide

Compound A-12 (15 mg, 0.025 mmol), HATU (9.3 mg, 0.025 mmol), DIEA (19mg, 0.15 mmol) and 3-methoxypropionic acid (3 mg, 0.029 mmol) werestirred at room temperature for 1 h, and then the mixture was pouredinto water (1 mL) and extracted with ethyl acetate (5 mL×2). The organicphase was washed with saturated aqueous NaCl solution, separated, driedover anhydrous Na₂SO₄, filtered and evaporated to dryness. The residuewas purified by column chromatography (DCM/MeOH=20:1) to give theproduct A-13-2 (6 mg, yield: 40%). LC-MS m/z=498.7 [M+1]+.

A-13-3: N-(4-(8-amino-1-(4-phenoxyphenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)-3-methyloxetane-3-carboxamide

A solution of compound A-12 (15 mg, 0.025 mmol), HATU (9.3 mg, 0.025mmol), DIEA (19 mg, 0.15 mmol) and 3-methyloxetane-3-carboxylic acid(3.4 mg, 0.029 mmol) in DMF (1 mL) was stirred at room temperature for 1h, and then the mixture was poured into water (5 mL) and extracted withethyl acetate (5 mL 2). The organic phase was washed with saturatedaqueous NaCl solution, separated, dried over anhydrous Na₂SO₄, filteredand evaporated to dryness. The residue was purified by columnchromatography (DCM/MeOH=20:1) to give the product A-13-3 (8 mg, yield:53%). LC-MS m/z=510.2 [M+1]+.

A-13-4:N-(4-(8-amino-1-(4-phenoxyphenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)-2-morpholinoacetamide

A solution of compound A-12 (15 mg, 0.025 mmol), HATU (9.3 mg, 0.025mmol), DIEA (19 mg, 0.15 mmol) and 2-morpholinoacetic acid (4.2 mg,0.029 mmol) in DMF (1 mL) was stirred at room temperature for 1 h, andthen the mixture was poured into water (1 mL) and extracted with ethylacetate (5 mL×2). The organic phase was wished with saturated aqueousNaCl solution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product A-13-4 (9 mg, yield: 58%).

Synthetic Route of Compound A-16-n

A-14: benzyl(4-(8-amino-1-(4-((5-fluoro-2-methoxybenzamido)methyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

A solution of benzyl(4-(8-amino-1-iodoimidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)carbamate(A-4) (300 mg, 0.6 mmol),5-fluoro-2-methoxy-N-(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl)benzamide(II-12) (284 mg, 0.738 mmol), Pd[PPh₃]₄ (69 mg, 0.06 mmol) and Cs₂O₃(239 mg, 0.738 mmol) in a mixed solvent of DME:H₂O (2.5 mL:0.5 mL) washeated to 80° C. and stirred overnight. The mixture was thenconcentrated, and the residue was purified by column chromatography togive the desired compound A-14 (285 mg, yield: 75%).

A-15:N-(4-(8-amino-3-(4-aminobicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)benzyl)-5-fluoro-2-methoxybenzamide

A solution of compound A-14 (280 mg, 0.44 mmol) in a mixed solvent ofDCM/TFA (10 mL:10 mL) was heated to 60° C. and stirred for 18 h. Afterthe mixture was evaporated to dryness, DCM (20 mL×2) was added, and theresulting mixture was concentrated. The residue was dissolved in DCM (30mL), and then the solution was added to a solution of HCl in dioxane (10mL). After the mixture was evaporated to dryness, DCM (20 mL×2) was thenadded, and the resulting mixture was concentrated, followed by theaddition of isopropyl ether (30 mL). After being stirred for 2 h, themixture was filtered and washed with isopropyl ether (10 mL×2) to givethe hydrochloride of the desired product A-15 (180 mg), which wasdirectly used in the next step without purification.

A-16-1:N-(4-(8-amino-3-(4-(2-hydroxy-2-methylpropanamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)benzyl)-5-fluoro-2-methoxybenzamide

A solution of compound A-15 (15 mg, 0.025 mmol), HATU (9.3 mg, 0.025mmol), DIEA (19 mg, 0.15 mmol) and 2-hydroxy-2-propionic acid (2.6 mg,0.025 mmol) in DMF (1 mL) was stirred at room temperature for 1 h, andthen the mixture was poured into water (1 mL) and extracted with ethylacetate (5 mL×2). The organic phase was washed with saturated aqueousNaCl solution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product A-16-1 (6 mg, yield: 40%). LC-MSm/z=587.3 [M+1]+.

A-16-2: N-(4(8-amino-3-(4-(3-methoxypropanamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)benzyl)-5-fluoro-2-methoxybenzamide

A solution of compound A-15 (15 mg, 0.025 mmol), HATU (9.3 mg, 0.025mmol), DIEA (19 mg, 0.15 mmol) and 3-methoxypropionic acid (2.6 mg,0.025 mmol) in DMF (1 mL) was stirred at room temperature for 1 h, andthen the mixture was poured into water (1 mL) and extracted with ethylacetate (5 mL×2). The organic phase was washed with saturated aqueousNaCl solution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product A-16-2 (8 mg, yield: 40%). LC-MSm/z=587.3 [M+1]+.

A-16-3:N-(4-(8-amino-1-(4-((5-fluoro-2-methoxybenzamido)methyl)phenyl)imidazo[1,5-a]pyrazin-3-yl)bicyclo[2.2.1]heptan-1-yl)-3-methyloxetane-3-carboxamide

A solution of compound A-15 (15 mg, 0.025 mmol), HATU (9.3 mg, 0.025mmol), DIEA (19 mg, 0.15 mmol) and 3-methyloxetane-3-carboxylic acid(2.9 mg, 0.025 mmol) in DMF (1 mL) was stirred at room temperature for 1h, and then the mixture was poured into water (5 mL) and extracted withethyl acetate (5 mL×2). The organic phase was washed with saturatedaqueous NaCl solution, separated, dried over anhydrous Na₂SO₄, filteredand evaporated to dryness. The residue was purified by columnchromatography (DCM/MeOH=20:1) to give the product A-16-3 (10 mg, yield:66%). LC-MS m/z=599.3 [M+1]+.

A-16-4:N-(4-(8-amino-3-(4-(2-morpholinoacetamido)bicyclo[2.2.1]heptan-1-yl)imidazo[1,5-a]pyrazin-1-yl)benzyl)-5-fluoro-2-methoxybenzamide

A solution of compound A-15 (15 mg, 0.025 mmol), HATU (9.3 mg, 0.025mmol), DIEA (19 mg, 0.15 mmol) and 2-morpholinoacetic acid (3.6 mg,0.025 mmol) in DMF (1 mL) was stirred at room temperature for 1 h, andthen the mixture was poured into water (1 mL) and extracted with ethylacetate (5 mL×2). The organic phase was washed with saturated aqueousNaCl solution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product A-16-4 (9 mg, yield: 57%). LC-MSm/z=628.3 [M+1]+.

Synthetic Route of Compound B-6-n

B-4: benzyl(4-(4-amino-5-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

A solution of compound B-3 (200 mg, 0.4 mmol),(4-((4-(trifluoromethyl)pyridin-2-yl) carbamoyl)phenyl)boronic acid(II-3)(152 mg, 0.49 mmol), Pd[PPh₃]₄ (46 mg, 0.04 mmol) and Cs₂CO₃ (159mg, 0.49 mmol) in a mixed solvent of DME:H₂O (2.5 mL:0.5 mL) was heatedto 80° C. and stirred overnight. The mixture was concentrated, and theresidue was purified by column chromatography (methanol/DCM=1:40) togive the desired compound B-4 (187 mg, yield: 73%).

B-5:4-(4-amino-7-(4-aminobicyclo[2.2.1]heptan-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-5-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound B-4 (187 mg, 0.29 mmol) in a mixed solvent ofDCM/TFA (10 mL:10 mL) was heated to 60° C. and stirred for 18 h. Afterthe mixture was evaporated to dryness, DCM (20 mL×2) was added, and theresulting mixture was concentrated. The residue was added with DCM (30mL), and then the mixture was added to a solution of HCl in dioxane (10mL). After the mixture was evaporated to dryness, DCM (20 mL×2) was thenadded again, and the resulting mixture was evaporated to dryness,followed by the addition of isopropyl ether (30 mL). After being stirredfor 2 h, the mixture was filtered and washed with isopropyl ether (10mL×2) to give the hydrochloride of the desired product B-5, which wasdirectly used in the next step without purification.

B-6-1:4-(4-amino-7-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-5-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound B-5 (19 mg, 0.031 mmol), HATU (12 mg, 0.031mmol), DIEA (24 mg, 0.19 mmol) and but-2-ynoic acid (2.6 mg, 0.031 mmol)in DMF (1 mL) was stirred at room temperature for 1 h, and then themixture was poured into water (1 mL) and extracted with ethyl acetate (5mL×2). The organic phase was washed with saturated aqueous NaClsolution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product B-6-1 (11 mg yield: 59%). LC-MSm/z=574.3 [M+1]+.

B-6-2:4-(4-amino-7-(4-(2-hydroxy-2-methylpropanamido)bicyclo[2.2.1]heptan-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-5-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound B-5 (19 mg, 0.031 mmol), HATU (12 mg, 0.031mmol), DIEA (24 mg, 0.19 mmol) and 2-hydroxy-2-methylpropionic acid (3.2mg, 0.031 mmol) in DMF (1 mL) was stirred at room temperature for 1 h,and then the mixture was poured into water (1 mL) and extracted withethyl acetate (5 mL×2). The organic phase was washed with saturatedaqueous NaCl solution, separated, dried over anhydrous Na₂SO₄, filteredand evaporated to dryness. The residue was purified by columnchromatography (DCM/MeOH=20:1) to give the product B-6-2 (9 mg, yield:49%). LC-MS m/t=594.2 [M+1]+.

B-6-3:4-(4-amino-7-(4-(3-methoxypropanamido)bicyclo[2.2.1]heptan-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-5-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound B-5 (19 mg, 0.031 mmol), HATU (12 mg, 0.031mmol), DIEA (24 mg, 0.19 mmol) and 3-methoxypropionic acid (3.2 mg,0.031 mmol) in DMF (1 mL) was stirred at room temperature for 1 h, andthen the mixture was poured into water (1 mL) and extracted with ethylacetate (5 mL×2). The organic phase was washed with saturated aqueousNaCl solution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product B-6-3 (12 mg, yield: 66%). LC-MSm/z=594.2 [M+1]+.

B-6-4:N-(4-(4-amino-5-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.1]heptan-1-yl)-3-methyloxetane-3-carboxamide

A solution of compound B-5 (19 mg, 0.031 mmol), HATU (12 mg, 0.031mmol), DIEA (24 mg, 0.19 mmol) and 3-methyloxetane-3-carboxylic acid(3.6 mg, 0.031 mmol) in DMF (1 mL) was stirred at room temperature for 1h, and then the mixture was poured into water (1 mL) and extracted withethyl acetate (5 mL×2). The organic phase was washed with saturatedaqueous NaCl solution, separated, dried over anhydrous Na₂SO₄, filteredand evaporated to dryness. The residue was purified by columnchromatography (DCM/MeOH=20:1) to give the product B-6-4 (12 mg, yield:64%). LC-MS m/z=606.3 [M+1]+.

B-6-5:4-(4-amino-7-(4-(2-morpholinoacetamido)bicyclo[2.2.1]heptan-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-5-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound B-5 (19 mg, 0.031 mmol), HATU (12 mg, 0.031mmol), DIEA (24 mg, 0.19 mmol) and 2-morpholinoacetic acid (4.5 mg,0.031 mmol) in DMF (1 mL) was stirred at room temperature for 1 h, andthen the mixture was poured into water (I mL) and extracted with ethylacetate (5 mL×2). The organic phase was washed with saturated aqueousNaCl solution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product B-6-5 (11 mg, yield: 58%). LC-MSm/z=635.2 [M+1]+.

Synthetic Route of Compound C-7-n

C-5: benzyl(4-(6-amino-8-oxo-7-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)-7,8-dihydro-9H-purin-9-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

A solution of compound C-4 (100 mg, 0.25 mmol),(4-((4-(trifluoromethyl)pyridin-2-yl) carbamoyl)phenyl)boronic acid(II-3) (160 mg, 0.5 mmol), copper acetate (64) mg, 0.3 mmol) and Et₃N(30 mg, 0.3 mmol) in DCM (10 mL) was stirred at room temperature for 24h, and then the mixture was poured into water (20 mL) and extracted withethyl acetate (10 mL×2). The organic phase was separated, dried overanhydrous Na₂SO₄, filtered and evaporated to dryness. The residue waspurified by column chromatography (EA/PE=1:1 EA) to give the desiredcompound C-5 (47 mg, yield: 29%). LC-MS m/z=657.3 [M−1]⁻.

C-6:4-(6-amino-9-(4-aminobicyclo[2.2.1]heptan-1-yl)-8-oxo-8,9-dihydro-7H-purin-7-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound C-5 (45 mg, 0.09 mmol) in a mixed solvent ofDCM/TFA (10 mL:10 mL) was heated to 60° C. and stirred for 18 h. Afterthe mixture was evaporated to dryness, DCM (20 mL 2) was added, and theresulting mixture was concentrated. The residue was dissolved in DCM (30mL), and then the solution was added to a solution of HCl in dioxane (10mL). After the mixture was evaporated to dryness, DCM (20 mL×2) was thenadded, and the resulting mixture was concentrated, followed by theaddition of isopropyl ether (30 mL). After being stirred for 2 h, themixture was filtered and washed with isopropyl ether (10 mL×2) to givethe hydrochloride of the desired product C-6, which was directly used inthe next step without purification.

C-7-1:4-(6-amino-9-(4-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)-8-oxo-8,9-dihydro-7H-purin-7-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound C-6 (13 mg, 0.022 mmol), HATU (9 mg, 0.022 mmol),DIEA (16.8 mg, 0.13 mmol) and but-2-ynoic acid (1.8 mg, 0.022 mmol) inDMF (1 mL) was stirred at room temperature for 1 h, and then the mixturewas poured into water (1 mL) and extracted with ethyl acetate (5 mL×2).The organic phase was washed with saturated aqueous NaCl solution,separated, dried over anhydrous Na₂SO₄, filtered and evaporated todryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product C-7-1 (5 mg, yield: 38%). LC-MSm/z=591.2 [M+1]+.

C-7-2:4-(6-amino-9-(4-(2-morpholinoacetamido)bicyclo[2.2.1]heptan-1-yl)-8-oxo-8,9-dihydro-7H-purin-7-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A solution of compound C-6 (13 mg, 0.022 mmol), HATU (9 mg, 0.022 mmol).DIEA (16.8 mg, 0.13 mmol) and 2-morpholinoacetic acid (3.2 mg, 0.022mmol) in DMF (1 mL) was stirred at room temperature for 1 h, and thenthe mixture was poured into water (1 mL) and extracted with ethylacetate (5 mL×2). The organic phase was washed with saturated aqueousNaCl solution, separated, dried over anhydrous Na₂SO₄, filtered andevaporated to dryness. The residue was purified by column chromatography(DCM/MeOH=20:1) to give the product C-7-2 (6 mg, yield: 42%). LC-MSm/z=652.2 [M+1]+.

Synthetic Route of Compound D-8-n

D-6: benzyl(4-(4-amino-3-(4-((4-(trifluoromethyl)pyridin-2-yl)carbamoyl)phenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)bicyclo[2.2.1]heptan-1-yl)carbamate

A mixed solution of D-5 (96 mg, 0.19 mmol), II-3 (73 mg, 0.234 mmol),Pd[PPh₃]₄ (22 mg, 0.019 mmol) and Cs₂CO₃ (76.4 mg, 0.234 mmol) inDME/H₂O (9 mL/1 ml) was purged with N₂ for 1 min to remove oxygen andthen heated to 80° C. in a sealed tube and stirred for 12 h. After themixture was cooled, the reaction was quenched with brine (20 mL) and EA(50 mL×2) was added for extraction. The organic phase was dried overanhydrous Na₂SO₄, filtered and concentrated. The residue was purified bysilica gel column chromatography (eluent: PE/EA=1:1) to give the desiredproduct D-6 (42 mg, 34.4%). LC-MS m/z=643.2 [M+1]+.

D-7:4-(4-amino-1-(4-aminobicyclo[2.2.1]heptan-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A mixture of D-6 (42 mg, 0.065 mmol) in TFA/DCM (1 mL/1 mL) was stirredat 60° C. for 12 h. After the mixture was cooled, the solvent wasremoved by evaporation. A solution of HCl in dioxane (2 mL) was thenadded, and the resulting mixture was stirred for 10 min andconcentrated. The residue was added with isopropyl ether (10 mL),stirred, and concentrated. The above procedure was repeated twice. Thesolid was filtered, washed with isopropyl ether and dried to give thehydrochloride of the desired product D-7 (30 mg, 75%).

D4-1:4-(4-amino-1-(44-(but-2-ynamido)bicyclo[2.2.1]heptan-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A mixed solution of D-7 (15 mg, 0.025 mmol), but-2-ynoic acid (2 mg,0.024 mmol), HATU (9.1 mg, 0.024 mmol) and DIEA (0.041 mL, 0.241 mmol)in DMF (1 mL) was stirred at room temperature for 4 h. The reactionsolution was diluted with EA (30 mL) and washed with brine, and theorganic phase was dried over anhydrous Na₂SO₄, filtered andconcentrated. The residue was purified by silica gel columnchromatography (eluent: PE/EA=1:1) to give the desired product D-8-1 (12mg, 86%). LC-MS m/z=575.1 [M+1]+.

D-8-2:4-(4-amino-1-(4-(2-morpholinoacetamido)bicyclo[2.2.1]heptan-1-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)-N-(4-(trifluoromethyl)pyridin-2-yl)benzamide

A mixed solution of D-7 (15 mg, 0.025 mmol), 2-morpholinoacetic acid(3.5 mg, 0.024 mmol), HATU (9.1 mg, 0.024 mmol) and DIEA (0.041 mL,0.241 mmol) in DMF (1 mL) was stirred at room temperature for 4 h. Thereaction solution was diluted with EA (30 mL) and washed with brine, andthe organic phase was dried over anhydrous Na₂SO₄, filtered andconcentrated. The residue was purified by silica gel columnchromatography (eluent: PE/EA=1:1) to give the desired product D-8-2(8.5 mg, 56%). LC-MS m/z=636.3 [M+1]+.

1. In Vitro Inhibitory Activity Against BTK(Wt)/BTK(C481S)(Determinationof IC₅₀ Value)

(1) Method

The substrate solution was prepared by adding the substrate poly(Glu,Tyr) sodium salt (Sigma Aldrich, St. Louis, Mo.) to the substratereaction buffer (20 mM Hepes (pH 7.5), 10 mM MgCl₂, 1 mM EGTA, 0.02%Brij35, 0.02 mg/mL BSA, 0.1 mM Na₃VO₄, 2 mM DTT and 1% DMSO)(finalsubstrate concentration in the reaction solution was 0.2 μM). Testcompound was formulated into stock solutions at 10 mM concentration with100% DMSO, and 3-fold serial dilution for 10 doses was performed in a384-well cyclic olefin copolymer LDV microplate. BTK(wt) or BTK(C481S)kinase (recombinant human full-length protein, histidine tag, expressedin insect cells, invitrogen, Carlsbad, Calif.) was added to thesubstrate solution and mixed gently (final BTK concentration in thereaction solution was 8 nM). Test compound in 100% DMSO was then addedto the kinase reaction mixture by acoustic liquid transfer technology(Echo 550; nanoliter range)(Labcyte Inc, Sunny vale, CA) and incubatedfor 20 min at room temperature. 33P-ATP (specific activity 10 μCi/μL)was added to the reaction mixture to initiate the reaction, followed byincubation at room temperature for 2 h. A small portion of the reactionmixture was dropped onto P-81 ion exchange filter paper (Whatman). Afterwashing the unbound phosphate from the filter paper with 0.75% phosphatebuffer (three times) and drying, the remaining radioactivity on thefilter paper was measured. The kinase activity was expressed as thepercentage of the remaining, kinase activity in the test sample to thekinase activity in the vehicle (dimethyl sulfoxide) blank control. IC₅₀values were calculated by curve fitting the data obtained using Prism(GraphPad Software).

(2) Results

Compared with the second-generation BTK inhibitor acalabrutinib, most ofthe compounds disclosed herein have stronger inhibition capabilityagainst the activity of wild-type BTK(wt) enzyme. More importantly, mostof the compounds exhibit more excellent inhibition activity againstBTK(C481S) mutants, and some even show inhibition activity of less than0.1 nM.

TABLE 1 Inhibition against activity of BTK (wt)/BTK (C481S) enzyme (A ≤0.1 nM; 0.1 nM < B ≤ 1 nM; 1 nM < C ≤ 10 nM; 10 nM < D ≤ 50 nM) CompoundIC₅₀ IC₅₀ number BTK (wt) BTK (C48IS) A-7-2  C B A-7-3  B B A-7-5  C BA-7-9  C B A-7-12  C B A-7-14  C B A-10-1  C B A-10-3  C B A-10-5  C AA-10-6  C A A-10-7  C B A-10-8  C A A-10-10 C A A-10-11 C B A-10-12 C BA-10-13 C A A-10-14 C B A-10-15 C B A-10-17 D C A-10-18 D C A-10-19 D CA-10-20 D C A-10-21 C C A-10-22 C C A-13-1  C C A-13-2  C B A-13-3  C CA-13-4  D C A-16-1  D C A-16-2  C C A-16-3  C C A-16-4  C C B-6-1  C CB-6-2  D C B-6-3  C C B-6-4  C C B-6-5  C C C-7-1  C C C-7-2  C BAcalabrutinlb 18.6 nM 815 nM

2. Experiment on In Vitro Tumor Cell Proliferation Inhibition

(1) Method

-   -   The tumor cell line (TMD-8/OCY-LY10) was suspended in        RPMI1640+FBS10% and cultured in an incubator (37° C. 5% CO₂).        Cells were passaged periodically, and cells at logarithmic        growth phase were collected for plating.    -   Cell staining was performed with trypan blue and viable cells        were counted.    -   Cell concentration was adjusted to 7000 cells/well with medium.    -   90 μL of cell suspension was added to a 96-well plate and        cell-free culture medium was added to the blank control wells.    -   Cells in the 96-well plate were incubated overnight in an        incubator (37° C., 5% CO₂, 100% relative humidity).    -   Preparation of 400× compound storage plates: test compounds and        reference drug were each dissolved in DMSO and 3× diluted from        the highest concentration (400 μM) to the lowest concentration        (0.61 μM).    -   Preparation of 10× compound working solutions: 78 μL of cell        culture medium was added to a V-bottom 96-well plate, and 2 μL        of the compound was pipetted from the 400× compound storage        plate into the 96-well plate. Vehicle control wells and blank        control wells were added with 2 μL of DMSO. After the addition        of the compound or DMSO, the mixture was mixed well by a        pipette.    -   Adding compound: 10 μL of 10× compound working solution (as        shown in Table 1) was added to the cell culture plate. 10 μL of        the mixture of DMSO and cell culture solution was added to the        vehicle control wells and blank control wells. The final        concentration of DMSO was 0.25%.    -   The 96-well cell plate was put back in the incubator for 72 h of        culturing.    -   CellTiter-Glo buffer was thawed and left to stand to room        temperature.    -   CellTiter-Glo substrate was left to stand to room temperature.    -   A flask of CellTiter-Glo substrate was added with the        CellTiter-Glo buffer for dissolving, such that a CellTiter-Glo        working solution was prepared.    -   The flask was subjected to slow vortex shaking to completely        dissolve the substrate.    -   The cell culture plate was taken out and left to stand for 30        min to be balanced to room temperature.    -   50 μL (equal to half the volume of cell culture medium in each        well) of the CellTiter-Glo working solution was added to each        well. The cell plate was wrapped with aluminum foil to keep out        of light.    -   The plate was shaken on at orbital shaker for 2 min to induce        cell lysis.    -   The plate was left to stand at room temperature for 10 min to        stabilize the luminescence signals.    -   The luminescence signals were detected on 2104 EnVision plate        reader.    -   Data analysis: The inhibition rate (IR) of the test compound was        calculated by the following formula: IR (%)=(1−(RLU compound        RLU−blank control)/(RLU vehicle control−RLU blank        control))×100%. Inhibition rates of compounds at different        concentrations were calculated in Excel, followed by plotting        the inhibition curves and calculating IC₅₀ values with GraphPad        Prism.

(2) Results

In the proliferation inhibition experiment of B-cell lymphoma cellsTMD-8 and OCI-LY10, compound A-10-10 and ibrutinib exhibit stronginhibition activity against tumor cell proliferation.

TABLE 2 Inhibitory activity olcompound A-10-10/ ibrutinib against tumorcell proliferation Compound IC₅₀ (nM) number TMD-8 OCI-LY10 A-10-10 2.910.3  Ibrutinib 0.4 1.7

3. Pharmacokinetic Study

(1) Method

Male SD rats were fasted overnight before test. The test drugs were eachsuspended in a solution of 0.5% methylcellulose (MC) and 0.1% SDS indeionized water (w/w/v), and the suspension, each at a concentration of1 mg/mL, was administered intragastrically at 5 mL/kg. At 15 min. 30min. 1 h, 2 h, 4 h, 6 h, 8 h and 24 h after administration,approximately 0.4 mL of whole blood was collected from the orbitalvenous plexus of the animals and then placed in heparin anticoagulanttubes. Three animals were sampled at each time point and then put todeath, and thus sample collection was completed on differentindividuals. The whole blood sample would be centrifuged within 15 minand centrifuged at 4° C./4200 rpm for 5 min. All plasma samples werestored in a refrigerator at −80±15° C. prior to analysis. An LC-MS/MS(Waters I Class UPLC-Xevo TQD tandem mass spectrometry) assay for thecompounds was established prior to sample analysis. The collected plasmawas quantitatively analyzed, plasma concentration-time data of animalswere analyzed using WinNonlin (professional edition, version 5.2)software, a non-compartmental model was used for concentration analysis,and the pharmacokinetic parameters of the test compounds werecalculated.

Three adult male beagle dogs were fasted overnight before test. The testdrugs were each suspended in a solution of 0.5% methylcellulose (MC) and0.1% SDS in deionized water (w/w/v), and the suspension, each at aconcentration of 1 mg/mL, was administered intragastrically at 5 mL/kg.Blood was collected 15 min, 30 min, 1 h, 2 h, 4 h, 6 h, 8 h and 24 hafter administration. Animals were lightly anesthetized with chloralhydrate, and approximately 0.5 mL of whole blood was collected from theforelimb vein using a glass blood collection tube and placed in aheparin anticoagulant tube. The sample was centrifuged at 4° C./4200 rpmfor 5 min. All plasma samples were stored in a refrigerator at −80±15°C. prior to analysis. An LC-MS/MS (Waters I Class UPLC-Xevo TQD tandemmass spectrometry) assay for the compounds was established prior tosample analysis. The collected plasma was quantitatively analyzed,plasma concentration-time data of animals were analyzed using WinNonlin(professional edition, version 5.2) software, a non-compartmental modelwas used for concentration analysis, and the pharmacokinetic parametersof the test compounds were calculated.

(2) Results

Compound A-10-10 (administered intragastrically, 5 mg/kg) showed goodabsorption with C_(max) of 8323 ng/mL and 641 ng/mL in rat and beagledog respectively, and it had a fairly high blood exposure(AUC_(0-INF)=73318 hr*ng/mL and 14867 hr*ng/mL).

TABLE 3 Pharmacokinetic data for rat/beagle (male) subjected tointragastric administration (5 mg/kg) (t_(1/2): half life; T_(max): peaktime; C_(max): maximum plasma concentration; AUC_(0-INF): area under0-INF time-concentration curve) Compound A-10-10 PK (p.o., 5 mg/kg)parameters Unit Rat Dog t_(1/2) hr 3.74 14.2 T_(max) hr 2.00 8.00C_(max) ng/mL 8323 641 AUC_(0-INF) hr*ng/mL 73318 14867

4. pBTK Inhibition Experiment on HEK293 Cells Transfected with BTK(Wt)and BTK(C481S)

The major cause of the drug resistance to ibrutinib lies in C481Smutation of BTK kinase, and developing a BTK inhibitor which caneffectively inhibit cells with BTK(C481S) variation is of greatsignificance for overcoming the drug resistance to ibrutinib.

(1) Method

-   -   HEK293 human embryonic kidney cells were transiently transfected        with human full-length BTK or BTK(C481S) vector.    -   Cells were dispensed into a 96-well plate at a predetermined        cell density.    -   Each test compound was subjected to 3-fold serial dilution for        eight times with 100% DMSO.    -   The compound was then diluted in tissue culture medium to        10-fold final assay concentration and 5% DMSO.    -   The compound was added to cells in the %-well plate (10-fold        dilution in culture medium) and the final concentration was        1×compound and 0.5% DMSO. For positive (high signal) controls,        cells were treated with 0.5% DMSO alone. For negative (low        signal) controls, cells were treated with 20 μM ibrutinib, and        the final concentration was 0.5% DMSO.    -   Cells were incubated with the compounds for 2 h at 37° C.    -   The cells were lysed and the lysate was transferred to an ELISA        plate previously coated with an antibody that captures the        substrate (human BTK or BTK(C481S)).    -   The plate was washed and then incubated with HRP-linked antibody        to detect total tyrosine phosphorylation.    -   The plate was washed and then added with HRP substrate. The        absorbance was read at 450 nm.    -   Based on the absorbance readings of the positive and negative        control values, the % inhibition value was calculated according        to the following formula: (% INH=((positive        control−sample)/(positive control−negative control))−100.    -   The Z value was calculated based on the following formula:        1−((3×standard deviation of positive+3×standard deviation of        negative)/(mean positive−mean negative)).    -   The logarithm of % inhibition value vs. compound concentration        was plotted using GraphPad Prism.    -   IC₅₀ values were determined after sigmoidal dose-response curve        fitting.

(2) Results

The C481S mutation reduces the inhibition of ibrutinib against BTKphosphorylation of HEK293 cells from 0.021 μM to 1.58 μM, while thecompound A-10-10 disclosed herein has strong inhibition (0.077 μM)against HEK293 cells transfected with BTK(wt) and even strongerinhibition (0.066 μM) against HEK293 cells transfected with BTK(C481S).

TABLE 4 Inhibition values of pBTK (IC₅₀) against HEK293 cellstransfected with BTK (wt) and BTK (C481S) Transfected in HEK293 cellsBTK (wt) BTK (C481S) pBTK IC₅₀ pBTK IC₅₀ Inhibitors (μM) (μM) A-10-100.077  0.063 Ibrutinib 0.021  1.58

5. Pharmacodynamic Experiment 1

(1) Method

CB17/SCID female mice with serious immune deficiency were purchased fromBeijing Vital River Laboratory Animal Technology Co., Ltd, and bred inSPF animal houses. Human OCI-LY10 cells (Shanghai Junrui-UFBN0102) werecultured in an RPMI 1640 culture medium containing 10% fetal bovineserum, 100 U/mL penicillin and 100 μg/mL streptomycin through monolayerculture in vitro in an incubator (37° C. 5% CO₂). Routine treatment wasperformed twice a week for passaging. At a cell saturation of 80%-90%and a required number, the cells were harvested and counted. 0.2 mL(1×10⁷) of OCI-LY10 cells (together with matrigel at a volume ratio of1:1) were subcutaneously inoculated into the right back of each mouse,and the tumor size could be measured about one week after inoculation.Tumor size was measured using a vernier caliper and tumor volume wascalculated using the following formula: tumor volume=(length×width²)/2.When the mean tumor volume reached 109 mm³, the mice were divided intofour groups (8 mice per group) for administration. i.e., vehicle (5%DMSO+20% HP-β-CD) control group, ibrutinib intragastric administrationgroup (25 mg/kg, once/day), and compound A-10-10 intragastricadministration group (25 mg/kg, 50 mg/kg, twice/day). Ibrutinib orcompound A-10-10 was dissolved in 5% DMSO+20% HP-β-CD and administeredintragastrically at 10 mL/kg for 28 consecutive days.

(2) Results

Compound A-10-10 and ibrutinib showed extremely strong anti-tumoractivity in OCI-LY10 xenograft tumor model (FIG. 1). The compoundA-10-10 intragastrically administered (the dosage was 25 mg/kg, bid; 50mg/kg, bid) could significantly inhibit the growth of the diffuse largeB-cell lymphoma cell strain OCI-LY10 (TG=110%, 110%; p<0.001, p<0.001).After 21 days of administration, all OCI-LY10 xenograft tumorscompletely disappeared in the 50 mg/kg group. After 28 days ofadministration, all OCI-LY10 xenograft tumors also disappearedcompletely in the 25 mg/kg group. At day 28, the TGI value for thecontrol compound ibrutinib 25 mg/kg group was 94% (p<0.001).

The effect of the test substance on body weight of tumor-bearing mice isshown in FIG. 1. The tumor-bearing mice showed good tolerance to thetest drug A-10-10 at all dosages, and no significant weight loss wasobserved in all treatment groups.

6. Pharmacodynamic Experiment 2

(1) Method

CB17/SCID female mice with serious immune deficiency were purchased fromBeijing Vital River Laboratory Animal Technology Co., Ltd, and bred inSPF animal houses. Human TMD-8 cells (Shanghai Junrui-UFBN1682) werecultured in an RPMI 1640 culture medium containing 10% fetal bovineserum, 100 U/mL penicillin and 100 μg/mL streptomycin through monolayerculture in vitro in an incubator (37° C., 5% CO₂). Routine treatment wasperformed twice a week for passaging. At a cell saturation of 80%-90%and a required number, the cells were harvested and counted. 0.2 mL(1×10⁷) of TMD-8 cells (together with matrigel at a volume ratio of 1:1)were subcutaneously inoculated into the right back of each mouse, andthe tumor size could be measured about one week after inoculation. Tumorsize was measured using a vernier caliper and tumor volume wascalculated using the following formula: tumor volume (length×width²)/2.When the mean tumor volume reached 107 mm³, the mice were divided intofour groups (8 mice per group) for administration, i.e., vehicle (5%DMSO+20% HP-β-CD) control group, ibrutinib intragastric administrationgroup (25 mg/kg, once/day), and compound A-10-10 intragastricadministration group (25 mg/kg, 50 mg/kg, twice/day). Ibrutinib orcompound A-10-10 was dissolved in 5% DMSO+20% HP-β-CD and administeredintragastrically at 10 mL % kg for 27 consecutive days.

(2) Results

Compound A-10-10 and ibrutinib showed extremely strong anti-tumoractivity in TMD-8 xenograft tumor model (FIG. 2). The compound A-10-10intragastrically administered (the dosage was 25 mg/kg, bid; 50 mg/kg,bid) could significantly inhibit the growth of the diffuse large B-celllymphoma cell strain TMD-8 (TGI=94%, 104%; p<0.001, p<0.001). After 27days of administration, 5/8 of the TMD-8 xenograft tumors disappeared inthe 50 mg/kg group. The TGI value for the control compound ibrutinib 25mg/kg group was 90% (p<0.001).

The effect of the test substance on body weight of tumor-bearing mice isshown in FIG. 2. The tumor-bearing mice showed good tolerance to thetest drug A-10-10 at all dosages, and no significant weight loss wasobserved in all treatment groups.

What is claimed is:
 1. A compound of formula I

or a pharmaceutically acceptable salt thereof, wherein ring A is thefollowing structure:

ring B is the following structure:

ring C is selected from one of the following structures:

L is one of the following structures:

R¹ is the following structure:

wherein R³ is selected from substituted or unsubstituted C₁₋₆ alkyl,substituted or unsubstituted C₆₋₁₀ aryl, substituted or unsubstitutedC₁₋₉ heteroaryl, substituted or unsubstituted C₃₋₇ cycloalkyl,substituted or unsubstituted C₂₋₇ heterocycloalkyl; wherein for R³, asubstituent of the substituted C₁₋₆ alkyl, the substituted C₆₋₁₀ aryl,the substituted C₁₋₉ heteroaryl, the substituted C₃₋₇ cycloalkyl, thesubstituted C₂₋₇ heterocycloalkyl is selected from one or more of cyano,hydroxyl, C₁₋₄ alkyl, C₁₋₃ alkoxy, C₃₋₇ heterocycloalkyl; and R² is H.2.-9. (canceled)
 10. The compound according to claim 1, wherein R¹ isselected from one of the following structures:


11. (canceled)
 12. A compound selected from any one of the followingstructures:

13.-17. (canceled)
 18. A pharmaceutical composition comprising thecompound according to claim
 1. 19. A pharmaceutical formulationcomprising a therapeutically effective amount of the compound accordingto claim 1, and a pharmaceutically acceptable excipient. 20.-22.(canceled)